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  • Saccharomyces cerevisiae  (169)
  • Nitrogen fixation  (142)
  • Textbook of geodesy
  • Springer  (315)
  • Irkutsk : Ross. Akad. Nauk, Sibirskoe Otd., Inst. Zemnoj Kory
  • Krefeld : Geologischer Dienst Nordhein-Westfalen
  • 2005-2009  (3)
  • 1985-1989  (312)
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  • 1
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    Springer
    In:  Corporate, Florida, Springer, vol. Developments in Petroleum Science vol. 15B, no. Publ. No. 12, pp. 9, (3-540-24165-5, XXVI + 228 p.)
    Publication Date: 2006
    Description: This book provides a summary of geodynamic results from Iceland that presently are found in a great number of scientific articles, but have not been collected before in a book
    Keywords: Textbook of geodesy ; Crustal deformation (cf. Earthquake precursor: deformation or strain) ; Geodesy ; Plate tectonics ; GeodesyY
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  • 2
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    In:  Cambridge, Springer, vol. LXXVIII, no. 2, pp. 125-169, (ISBN: 3-540-42642-6, Approx. 620 p. 30 illus., Hardcover)
    Publication Date: 2006
    Keywords: Crustal deformation (cf. Earthquake precursor: deformation or strain) ; Volcanology ; Geodesy ; Global Positioning System ; InSAR ; Textbook of geodesy
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  • 3
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    In:  New York, 390 pp., Springer, vol. 36, no. XVI:, pp. 1-14, (ISBN 3-211-23584-1)
    Publication Date: 2005
    Description: Motivation, Fundamentals of potential theory The gravity field of the earth, Gravity reduction, Heights The geometry of the earth, Gravity field outside the earth, Space methods Modern views on the determination of the figure of the earth Statistical methods in physical geodesy, Least-squares collocation Computational methods, References, Subject index
    Keywords: Textbook of geodesy ; Global Positioning System ; Geoid ; Gravimetry ; Potential ; Theory ; Reference ; Systems ; collocation ; geodetic ; satellite ; techniques ; heights
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  • 4
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    Journal of industrial microbiology and biotechnology 4 (1989), S. 49-53 
    ISSN: 1476-5535
    Keywords: l-Phenylacetyl carbinol ; Saccharomyces cerevisiae ; Yeast ; Benzaldehyde ; Biotransformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The rate of production ofl-phenylacetyl carbinol bySaccharomyces cerevisiae in reaction mixtures containing benzaldehyde with sucrose or pyruvate as cosubstrate was investigated in short 1 h incubations. The effect of yeast dose rate, sucrose and benzaldehyde concentration and pH on the rate of reaction was determined. Maximum biotransformation rates were obtained with concentrations of benzaldehyde, sucrose and yeast of 6 g, 40 g and 60 g/l, respectively. Negligible biotransformation rates were observed at a concentration of 8 g/l benzaldehyde. The reaction had a pH optimum of 4.0–4.5. Rates of bioconversion of benzaldehyde and selected substituted aromatic aldehydes using both sucrose and sodium pyruvate as cosubstrate were compared. The rate of aromatic alcohol production was much higher when sucrose was used rather than pyruvate.o-Tolualdehyde and 1-chlorobenzaldehyde were poor substrates for aromatic carbinol formation although the latter produced significant aromatic alcohol in sucrose-containing media. Yields of 2.74 and 3.80 g/l phenylacetyl carbinol were produced from sucrose and pyruvate, respectively, in a 1 h reaction period.
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  • 5
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    Journal of industrial microbiology and biotechnology 4 (1989), S. 81-84 
    ISSN: 1476-5535
    Keywords: Ethanol fermentation ; Wheat starch ; Saccharomyces cerevisiae ; immobilization ; Continuous dynamic immobilized biocatalyst bioreactor ; Biocatalyst bioreactor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary A simple and efficient method of conversion of wheat starch B to ethanol was investigated. Employing a two-stage enzymatic saccharification process, 95% of the wheat starch was converted to fermentable sugars in 40 h. From 140 g/l total sugars in the feed solution, 63.6 g/l ethanol was produced continuously with a residence time of 3.3 h in a continuous dynamic immobilized biocatalyst bioreactor by immobilized cells ofSaccharomyces cerevisiae. The advantages and the application of this bioreactor to continuous alcoholic fermentation of industrial substrates are presented.
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  • 6
    ISSN: 1572-8773
    Keywords: Manganese ; Electron spin resonance ; Superoxide dismutase ; Saccharomyces cerevisiae ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary Manganese accumulation was studied by room-temperature electron spin resonance (ESR) spectroscopy inSaccharomyces cerevisiae grown in the presence of increasing amounts of MnSO4. Mn2+ retention was nearly linear in intact cells for fractions related to both low-molecular-mass and macromolecular complexes (‘free’ and ‘bound’ Mn2+, respectively). A deviation from linearity was observed in cell extracts between the control value and 0.1 mM Mn2+, indicating more efficient accumulation at low Mn2+ concentrations. The difference in slopes between the two straight lines describing Mn2+ retention at concentrations lower and higher than 0.1 mM, respectively, was quite large for the free Mn2+ fraction. Furthermore it was unaffected by subsequent dialyses of the extracts, showing stable retention in the form of low-molecular-mass complexes. In contrast, the slope of the line describing retention of ‘bound’ Mn2+ at concentrations higher than 0.1 mM became less steep after subsequent dialyses of the cell extracts. This result indicates that the macromolecule-bound Mn2+ was essentially associated with particulate structures. In contrast to Cu2+, Mn2+ had no effect on the major enzyme activities involved in oxygen metabolism except for a slight increase of cyanide-resistant Mn-superoxide dismutase activity, due to dialyzable Mn2+ complexes.
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  • 7
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    Biology and fertility of soils 8 (1989), S. 356-368 
    ISSN: 1432-0789
    Keywords: Plant-root associations ; Azospirillum spp ; Rhizosphere ; Nitrogen fixation ; Acetylene reduction assay (ARA) ; Phytohormones
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Bacteria of the genus Azospirillum are extensively studied for their plant-growth promoting effect following inoculation. Physiological and biochemical studies of these diazotrophic bacteria are now benefiting from recent breakthroughs in the development of genetic tools for Azospirilum. Moreover, the identification and cloning of Azospirillum genes involved in N2 fixation, plant interaction, and phytohormone production have given new life to many research projects on Azospirillum. The finding that Azospirillum genes can complement specific mutations in other intensively studied rhizosphere bacteria like Rhizobia will certainly trigger the exploration of new areas in rhizosphere biology. Therefore a review of the Azospirillum-plant interactions is particularly timely.
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  • 8
    ISSN: 1432-0789
    Keywords: Nodule damage ; Rivellia angulata ; Nitrogen fixation ; Cajanus cajan ; Pigeonpea ; Vertisol
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Damage caused by Rivellia angulata larvae to pigeonpea root nodules at the ICRISAT center in India was greater in the crop grown on Vertisols (up to 86%) compared to that on Alfisols (20%). Attempts to quantify the field effects of nodule damage on growth and yield of pigeonpea in a Vertisol, involving many heavy applications of soil insecticides (aldrin and hexachlorocyclohexane) failed because the insecticides did not control the pest and adversely affected the growth of the pigeonpea and the subsequent crop of sorghum (Sorgorum bicolor L. Moench). The impact of nodule damage on pigeonpea growth, yield and nutrient uptake was successfully studied in greenhouse-grown plants at three N levels. In this pot study, artificial inoculation with Rivellia sp. led to substantial nodule damage (70%). The results of this damage were a significant overall reduction in nodule dry weight (46%), acetylene reduction activity (31%), total leaf area (36%), chlorophyll content of leaves (39%) and shoot dry weight (23%) 68 days after sowing. At maturity, Rivellia sp. infestation caused significant reductions in top dry weight (22%), root and nodule dry weight (27%), seed dry weight (14%), and total N (29%) and P uptake (19%). The problems and prospects of manipulating nodule damage so as to reduce N losses in pigeonpea are discussed.
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  • 9
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    Biology and fertility of soils 7 (1989), S. 269-274 
    ISSN: 1432-0789
    Keywords: Nitrogen fixation ; Frankia-Ceanothus spp. association ; Acetylene reduction assay (ARA) ; Microsymbiont population ; Nodules ; Actinomycetes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Wildland shrub improvement is needed for sound range and disturbed land revegetation practice. The possibility of selecting superior N2-fixingFrankia-Ceanothus spp. actinorhizal associations was examined. Greenhouse tests were used to expose various soil-borne microsymbiont andCeanothus sp. population accessions in reciprocal combination. The acetylene reduction rate was used as a measure of N2-fixation capacity. There was no significant interaction between host and microsymbiont regardless of source for all variables measured. The acetylene reduction rate, nodule number and mass, plant biomass, and root: shoot ratio were significantly different among soil sources. The acetylene reduction rate was not significantly different amongCeanothus sp. accessions. Neither was it strongly correlated with other variables. It was concluded that the N2-fixation rate is more a function ofFrankia sp. than the hostCeanothus sp. in actinorhizal associations. It appears possible to select soil sources with superior N2-fixing microsymbiont populations.
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  • 10
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Vector ; Glyphosate resistance ; Transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The E. coli aroA gene was inserted between yeast promoter and terminator sequences in different shuttle expression plasmids and found to confer enhanced EPSP synthase activity as well as resistance to glyphosate toxicity. Subsequently, a transformation system using these newly constructed vectors in yeast was characterized. The efficiency of the glyphosate resistance marker for transformation and selection with plasmid pHR6/20-1 in S. cerevisiae laboratory strain SHY2 was found to be relatively high when compared with selection for LEU2 prototrophy. The fate of the recombinant plasmid pHR6/20-1 in the transformants, the preservation of the aroA E. coli DNA fragment in yeast, mitotic stability, EPSP synthase activity, and growth on glyphosate-containing medium have been investigated. As this plasmid also allows direct selection for glyphosate resistant transformants on rich media, the glyphosate resistance marker was used for transforming both S. cerevisiae laboratory strain SHY2 and brewer's yeast strains S. cerevisiae var. “uvarum” BHS5 and BHS2. In all cases, the vector pHR6/20-1 was maintained as an autonomously replicating plasmid. The resistance marker is, therefore, suitable for transforming genetically unlabeled S. cerevisiae laboratory, wild, and industrial yeast strains.
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  • 11
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Chromosome V ; Ty elements ; tRNA genes ; Transposition hot-spots ; Yeast polymorphism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Ty4 is a novel transposable element in the yeast, Saccharomyces cerevisiae, which is present in only a few copies in the genome (Stucka et al. 1989). In strain C836 one of the three copies (Ty4-90) is contained in cosmid clone c90, where it resides on chromosome V. Analysis of this region reveals a “hot-spot” of transposition: in addition to Ty4-90, the locus contains a complete Ty3 element and seven singular delta, sigma and tau elements. Three tRNA genes (for His, Lys, and Ile) are located in this region, and these are closely associated with one or the other of the elements, a phenomenon commonly observed in yeast. A comparison of c90 with corresponding regions from other strains shows that the locus is highly polymorphic and that this polymorphism is explicitly associated with Ty transposition and recombination events.
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  • 12
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    Current genetics 16 (1989), S. 315-321 
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Antisuppressor ; ADE3 ; Translation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Mutations in a known yeast gene, ADE3, were shown to act as an antisuppressor, reducing the efficiency of the omnipotent suppressor, sup45-2. The ADE3 locus encodes the trifunctional enzyme C1-tetrahydrofolate synthase, which is required for the biosynthesis of purines, thymidylate, methionine, histidine, pantothenic acid and formylmethionyl-tRNAfmet. The role of this enzyme in translational fidelity had not previously been suspected.
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  • 13
    ISSN: 1432-0983
    Keywords: Yeast ; Saccharomyces cerevisiae ; Nonsense suppression ; Omnipotent suppressors ; Gene mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Ten dominant omnipotent suppressors of Saccharomyces cerevisiae, which were previously shown to be different from SUP46, have been examined. Nine are mapped in a region between lys5 and cyh2 on the left arm of chromosome VII. These suppressors, like SUP46, manifest sensitivity to increased temperature and the antibiotics paromomycin and hygromycin B. In addition, they have an identical action spectrum. These results strongly suggest that they are allelic to each other and they are designated SUP138. The tenth is mapped to a position between his1 and arg6 on the right arm of chromosome V. This suppressor, named SUP139, does not manifest temperature sensitivity nor antibiotic sensitivity. SUP139 and SUP138, which are clearly distinguished by means of action spectrum, act on much fewer nonsense mutations than SUP46. It is now clear that dominant omnipotent suppressors arising at a single locus are homogeneous and that their efficiency is locus-dependent. The order of efficiency is SUP46〉SUP138〉SUP139.
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  • 14
    ISSN: 1432-0983
    Keywords: Yeast ; Repair ; Complementation ; Saccharomyces cerevisiae ; Schizosaccharomyces pombe ; Gene cloning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two Saccharomyces cerevisiae genes necessary for excision repair of UV damage in DNA, RAD1 and RAD2, were introduced individually, on a yeast shuttle vector, into seven Schizosaccharomyces pombe mutants — rads1, 2, 5, 13, 15,16 and 17. The presence of the cloned RAD1 gene did not affect survival of any of the S. pombe mutants. The RAD2 gene increased survival of S. pombe rad13 to near the wild-type level after UV irradiation and had no effect on any of the other mutants tested. S. pombe rad13 mutants are somewhat defective in removal of pyrimidine dimers so complementation by the S. cerevisiae RAD2 gene suggests that the genes may code for equivalent proteins in the two yeasts.
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  • 15
    ISSN: 1432-0983
    Keywords: DNA transformation ; Saccharomyces cerevisiae ; Site-specific recombination ; 2μ DNA plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The 2μ DNA plasmid of the yeast Saccharomyces cerevisiae does not confer any known selectable phenotype to the host cell carrying it. Selection of cells transformed with purified 2μ DNA therefore cannot be achieved, and the intracellular presence of 2μ can only be assessed by molecular analysis of the DNA complement. In addition, 2μ alone does not replicate in bacterial hosts, thus rendering its amplification by conventional methods impossible. We have isolated a shuttle plasmid, pBH-2L, generated by in vivo sites-pecific recombination between the endogenous 2μ DNA plasmid and pRL, a pBR322 derivative containing the yeast LEU2 gene and one 2μ repeat sequence associated with the origin of replication. This new shuttle plasmid has the property, when transformed into yeast, of undergoing site-specific recombinational resolution between its two direct repeat sequences. This releases 2μ plasmid and pRL as individual molecules. The latter can undergo progressive mitotic loss during growth in nonselective medium, ultimately leaving leucine auxotrophic transformants that contain only 2μ DNA plasmid. This system can be utilized to introduce 2μ DNA alone into cells lacking it, thereby providing a novel means to study the biology and the molecular genetics of the plasmid and its potential practical applications as a vector.
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  • 16
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    Current genetics 15 (1989), S. 113-120 
    ISSN: 1432-0983
    Keywords: Calmodulin mutant ; Nuclear division ; Chromosome stability ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The coding region of a yeast calmodulin gene was fused to a galactose-inducible GAL1 promoter, and a conditional-lethal mutant of Saccharomyces cerevisiae, in which the expression of calmodulin was regulated by galactose, was constructed. The mutant grew normally in galactose medium, but in glucose medium, in which the promoter was repressed, it ceased growing after 12–15 h. The growth arrest was associated with a decrease in intracellular calmodulin levels: after 12h, no intracellular calmodulin protein was detectable. Analysis of the terminal phenotype showed that when the cell stopped growing, it had a bud, a nucleus after S-phase and a short mitotic spindle. Thus, the defect was mainly in nuclear division. Bud growth was partially inhibited in these cells: 27% of the cells stopped growing with a small bud. Furthermore, calmodulin-deficient cells showed elevated rates of chromosome loss, possibly as the result of a defect in the precise segregation of chromosomes.
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  • 17
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    Current genetics 15 (1989), S. 235-237 
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mitochondrial DNA ; Method of extraction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A rapid method for the extraction of yeast mitochondrial DNA (mtDNA) is described. In comparison with previous methods, it simplifies several steps, does not require either the isolation of mitochondria or phenol treatment and is less time consuming. This protocol gives a high yield of pure mtDNA (50–120 μg from a 100-ml culture), which can be directly used in various molecular applications: restriction enzyme digestion, electrophoresis, blotting, labeling, cloning and sequencing.
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  • 18
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mutagen hyperresistance ; Southern, Northern analysis ; Gene transplacement ; Transposon mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The genes SNQ and SFA confer hyperresistance to 4-NQO and FA when present on a multi-copy plasmid in yeast. Both are non-essential genes since transplacement of SNQ by a disrupted snq-0::LEU2 yielded stable and viable haploid integrants. Southern analysis revealed that SNQ and SFA are single-loci genes, and OFAGE analysis showed that they are located on chromosome XIII and IV, respectively. Northern blot analysis of SNQ and SFA revealed poly(A)+ RNA transcripts of 2 kb and 1.7 kb, respectively. Nuclease S 1 mapping showed SNQ to have a coding region of 1.6 kb and SFA, one of 1.3 kb. The 5′ coding regions were determined for both genes, while the 3′ end could only be determined for gene SNQ. Both genes do not appear to contain introns. The SFA locus was also mapped by transposon mutagenesis. Tn10-LUK integrants disrupted the SFA gene function at sites that were determined by subcloning to lie within the SFA transcription unit.
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  • 19
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; SKI3 ; SKI5 ; M1 dsRNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have identified a mutant strain of the yeast Saccharomyces cerevisiae which overproduces killer toxin. This strain contains a single mutation which fails to complement defects in both the SKI3 and SKI5 genes, while a cloned copy of this gene complements both the ski3 and ski5 defects. The level of secreted toxin from a cDNA based plasmid is not increased in a ski3 strain, showing that the overproduction phenotype is dependent upon an increased level of M1 dsRNA.
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  • 20
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Psoralen photoaddition ; Interstrand cross-link ; Repair deficiency ; Genotoxicity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two different UVA irradiation systems were initially biologically calibrated with two haploid yeast strains proficient and deficient, respectively, in nucleotide excision repair. The number of DNA lesions introduced into the cell's genome by the photoactivated bifunctional furocoumarin 8-MOP was then calculated by means of the applied UVA exposure doses. At LD37 the repair-proficient wild type had about 14 ICL and 34 furan-side monoadducts in its DNA, while doubly blocked repair mutant rad3-12 pso1-1 had 2 ICL and 3 monoadducts. Locus-specific reversion of lys1-1 followed two-hit kinetics in the repair-proficient wild type and one-hit kinetics in an excision-deficient rad2-20 mutant, as would be expected if ICL was the main type of mutagenic lesion in the wild type and monoadducts the main mutagenic lesion type in the excision-deficient strain. Quantitative comparison of 8-MOP + UVA-induced ICL with those induced by bifunctional mustard revealed the former to have a much higher genotoxicity.
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  • 21
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Protoplast fusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The percentage of hybrids formed during protoplast fusion in Saccharomyces cerevisiae is determined by the percentage of protoplasts at the GI/S boundary of the cell cycle.
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  • 22
    ISSN: 1432-0983
    Keywords: Peroxisomes ; Protein import ; Saccharomyces cerevisiae ; Hansenula polymorpha
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The DAS gene of Hansenula polymorpha was expressed in Saccharomyces cerevisiae under the control of different promoters. The heterologously synthesized dihydroxyacetone synthase (DHAS), a peroxisomal enzyme in H. polymorpha, shows enzymatic activity in baker's yeast. The enzyme was imported into the peroxisomes of S. cerevisiae not only under the appropriate physiological conditions for peroxisome proliferation (oleic acid media), but also in glucose-grown cells where it induced the enlargement of the few peroxisomes present. This growth process was not accompanied by an increase in the number of microbodies, which suggests a separate control mechanism for peroxisomal proliferation.
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  • 23
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    Current genetics 15 (1989), S. 399-401 
    ISSN: 1432-0983
    Keywords: Saccharomyces exiguus ; Saccharomyces cerevisiae ; HO gene ; MAT gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The DNA of Saccharomyces exiguus was analyzed by Southern hybridization using cloned MATa, MATα, and HO genes of Saccharomyces cerevisiae as probes. It was shown that S. exiguus has a DNA sequence homologous with the HO gene of S. cerevisiae and that this DNA sequence is on a chromosome of about 940 kb of DNA in S. exiguus. However, there is no DNA sequence in S. exiguus that is homologous with the MAT genes of S. cerevisiae.
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  • 24
    ISSN: 1420-9071
    Keywords: Saccharomyces cerevisiae ; growth conditions ; kinaseless mutant ; plasma membrane vesicles ; glucose transport ; kinetics and computer simulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary In this study experimental data on the kinetic parameters investigated by other authors1–5, 11 together with own data on plasma membrane vesicles, have been subjected to a computer simulation based on the equations describing facilitated diffusion. The simulation led to an ideal fit describing the above data. From this it can be concluded that glucose is transported by facilitated diffusion, and not by active transport as was postulated by Van Steveninck14, 15. The simulation method also demonstrates that the fast sampling technique used by these authors1–5,11 underestimates the fluxes. Thus, the parameters given do not contribute to the understanding of glucose transport under different metabolic conditions. The K value of plasma membrane vesicles prepared from glucose-repressed cells is around 7 mM. Derepression, particularly by galactose, causes a highly significant increase in affinity as shown by a decrease in the K value to 2 mM. The highest affinity was measured in a triple kinaseless mutant grown on glycerol with a K value of 1 mM. If seems, therefore, that the kinetic parameters derived from initial uptake rates of glucose in intact cells1–5,11 using single flux analysis, such as Eadie-Hofstee- or Lineweaver-Burk-plots, are in error.
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  • 25
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    Oecologia 79 (1989), S. 566-568 
    ISSN: 1432-1939
    Keywords: Carbon dioxide ; Lichen ; Lobaria ; Nitrogen fixation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Thalli of Lobaria pulmonaria (L.) Hoffm., a nitrogen-fixing epiphyte common in mesic temperate forests, were collected in a Douglas-fir (Pseudotsuga menziesii Franco) forest near Corvallis, Oregon, and maintained for 20 to 40 days in controlled-environment chambers with atmospheric CO2 concentrations of 374 and 700 μll-1. Nitrogenase activity, which was assayed by the acetylene reduction method, was approximately doubled in the lichen maintained in elevated CO2. Increases in nitrogen fixation by lichens may be an important part of the integrated ecosystem response to rising CO2.
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  • 26
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Yeast ; Phospholipase B ; Lysophospholipase ; Enzyme inhibition ; AMP ; Unesterified fatty acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Divalent cations activate the lysophospholipase and transacylase reactions catalyzed by the same enzymes in the yeast Saccharomyces cerevisiae. The activation was observed at neutral pH, but not at the pH optimum of lysophospholipase/transacylase, near 3.5. Adenine nucleotides, especially AMP and ADP, are strong inhibitors of the same group of enzymes. Half maximal inhibition by AMP was found at a concentration of about 20 μM. The inhibition by nucleotides in low concentrations is enhanced by divalent cations.
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  • 27
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    Archives of microbiology 151 (1989), S. 445-453 
    ISSN: 1432-072X
    Keywords: Denitrification ; Growth yield measurements ; Nitrate respiration ; Nitrogen fixation ; Proton translocations in respirations ; Azospirillum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract For Azospirillum brasilense Sp7, the energy transformation efficiencies were measured in anaerobic respirations with either nitrate, nitrite or nitrous oxide as respiratory electron acceptors by determining the maximal molar growth yields and the H+-translocations using the oxidant pulse method. In continuous cultures grown with malate limiting, the maximal molar growth yields (Y s max -values) were essentially the same with O2 or N2O but were 1/3 and 2/3 lower with NO 2 - or NO 3 - , respectively, as respiratory electron acceptors. Both the maximal molar growth yields and the maintenance energy coefficients were surprisingly high when Azospirillum was grown with nitrite as the sole electron acceptor and source for N-assimilation. Growth under N2-fixing conditions drastically reduced the Y s max -values in the N2O and O2-respiring cells. In the H+-translocation measurements, the $$\vec H^ + $$ /oxidant ratios were 5.6 for O2→H2O, 2.5–2.8 for NO 3 - →NO 2 - , 2.2 for NO 2 - →N2O and 3.1 for N2O→N2 respirations when the cells were preincubated with valinomycin and K+. All the values were enhanced when the experiments were performed with valinomycin plus methyltriphenylphosphonium (=TPMP+) cation. The uncoupler carbonyl cyanide-m-chlorophenyl-hydrazone diminished the H+-excretion indicating that this translocation was due to vectorial flow across the membrane. In the absence of any ionophore, nitrate and nitrite respirations were accompanied by a H+-uptake $$(NO_3^ - \to N_2 = - 2.9 \vec H^ + /NO_3^ - and NO_2^ - \to N_2 = - 2.5 \vec H^ + /NO_2^ - )$$ . Any significant H+-translocation could not be detected in N2O- and O2-respirations under these conditions. It is concluded that nitrate reduction proceeds inside the cytoplasmic membrane, whereas nitrite is reduced extramembraneously. The data are not conclusive for the location of nitrous oxide reductase. The maximal molar growth yield determinations and the absence of any H+-uptake in untreated cells indicate a cytoplasmic orientation of the enzyme similar to the terminal cytochrome oxidase of respiration. The low H+-extrusion values for N2O-respiration compared to O2-respiration in cells treated with valinomycin plus TPMP+ are, however, not in accord with such an interpretation.
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  • 28
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    Archives of microbiology 151 (1989), S. 198-202 
    ISSN: 1432-072X
    Keywords: Sexual agglutination ; Mating ; Saccharomyces cerevisiae ; Yeast
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    Topics: Biology
    Notes: Abstract Genetic regulation of the inducibility of sexual agglutination ability in the yeast Saccharomyces cerevisiae was studied. Detailed analysis of the degree of sexual agglutination was carried out; it showed that a greater number of genes are involved in the regulation of inducible sexual agglutination in strain H1-0 than previously assumed. Although dominancy of inducible phenotype over constitutive was confirmed, the effectiveness of one gene changing the constitutive phenotype to the inducible seemed to be somewhat low. Quantity per cell of agglutination substances responsible for sexual agglutination increased as the agglutination ability became greater.
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  • 29
    ISSN: 1432-072X
    Keywords: Bradyrhizobium ; Gene cloning ; Heme ; Marker exchange mutagenesis ; Nitrogen fixation ; Respiration ; Symbiosis
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    Notes: Abstract Random and site-directed Tn5-induced mutagenesis of Bradyrhizobium japonicum yielded two mutations, one in strain 2960 and the other in strain 2606::Tn5-20, which mapped close to each other but in separate genes. The corresponding wild-type genes were cloned, and their approximate location on the cloned DNA was determined. Mutant 2960 was Fix- and formed green nodules on soybean, whereas strain 2606::Tn5-20 had ca. 4% of wild-type Fix activity and formed white nodules. Cytochrome oxidase assays (Nadi tests) showed a negative reaction with both mutants, indicating a functional deficiency of cytochrome c or its terminal oxidase or both. However, the mutants grew well under aerobic conditions on minimal media with different carbon sources. Furthermore, mutant 2960 had a reduced activity in hydrogen uptake, was unable to grow anaerobically with nitrate as the terminal electron acceptor and 2960-infected soybean nodules contained little, if any, functional leghemoglobin. Southern blot analysis showed that a B. japonicum heme biosynthesis mutant [strain LO505: O'Brian MR, Kirshbom PM, Maier RJ (1987) Proc Natl Acad Sci USA 84: 8390–8393] had its mutation close to the Tn5 insertion site of our mutant 2606::Tn5-20. This finding, combined with the observed phenotypes, suggested that the genes affected in mutants 2960 and 2606::Tn5-20 were involved in some steps of heme biosynthesis thus explaining the pleiotropic respiratory deficiencies of the mutants. Similar to strain LO505, the mutant 2606::Tn5-20 (but not 2960) was defective in the activity of protoporphyrinogen IX oxidase which catalyzes the penultimate step in the heme biosynthesis pathway. This suggests that one of the two cloned genes may code for this enzyme.
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  • 30
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    Archives of microbiology 151 (1989), S. 180-182 
    ISSN: 1432-072X
    Keywords: Klebsiella pneumoniae ; Nitrogen fixation ; nifL ; Regulation ; Oxygen control ; Nitrogen control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A number of in-frame deletions have been constructed in the Klebsiella pneumoniae regulatory gene nifL. The effects of each nifL mutation on NifA-mediated expression from the nifH promoter of K. pneumoniae have then been assessed with respect to both nitrogen and oxygen control. These experiments indicate that, in contrast to the situation with the homologous regulatory proteins NtrB and NtrC, NifA activity is not impaired in the absence of NifL. We conclude that the only function of NifL is to inactivate NifA in response to an increase in the nitrogen or oxygen status of the cell.
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  • 31
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    Archives of microbiology 151 (1989), S. 391-398 
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Exoglucanases ; Purification ; Protein moieties ; Tunicamycin
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    Topics: Biology
    Notes: Abstract Exoglucanase (exo-1,3-β-D-glucan glycohydrolase, EC 3.2.1.56) activity secreted by Saccharomyces cerevisiae into the culture medium was separated by ion exchange chromatography into two glycoprotein isoenzymes which contributed 10% (exoglucanase I) and 90% (exoglucanase II) towards the total activity. Analysis of the “in vitro” deglycosylated products by polyacrylamide gel electrophoresis under native or denaturing conditions indicated that the protein portions of both exoglucanases exhibited identical mobility, each one consisting of two polypeptides with M r of 47000 and 48000. The same profile was shown by the exoglucanase secreted in the presence of tunicamycin. Antibodies raised against the protein portion of exoglucanase II did react with both native exoglucanases and their deglycosylated products with a pattern indicative of immunological identity. Digestion of the “in vitro” deglycosylated products of both exoglucanases with Staphylococcus aureus V-8 protease or trypsin generated the same proteolytic fragments in each case. Only exoglucanase II was secreted by protoplasts. These and previously reported results indicate that the protein portions of both isoenzymes may be the product of the same gene (or a family of related genes), and that exoglucanase I is a product of enzyme II, modified by a process occurring beyond the permeability barrier of the cell.
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  • 32
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Recombination ; Tryptophan cluster ; Yeast vectors ; Plasmid copy number
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    Notes: Abstract We describe a convenient method for the in vivo construction of large plasmids that possess a multitude of restriction sites. A large (23 kbases) circular self-replicating plasmid carrying a partial LEU2-d gene was cotransformed with a circular non-replicating plasmid carrying the entire LEU2 gene. In vivo recombination results preferentially in a plasmid that carries both the LEU2-d and the entire LEU2 gene. In addition we also found one plasmid with a tandem LEU2 insertion and one plasmid where the LEU2-d gene was replaced by the entire LEU2 gene.
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  • 33
    ISSN: 1432-2048
    Keywords: Glutamate synthase ; Glutamine synthetase ; Nitrogen fixation ; Phaseolus (glutamate synthase) ; Plastid (glutamate synthase) ; Root nodule
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Abstract The two isoenzymes of NADH-dependent glutamate synthase (NADH-GOGAT; EC 1.4.1.14), previously identified in root nodules of Phaseolus vulgaris L., have both been shown to be located in root-nodule plastids. The nodule specific NADH-GOGAT II accounts for the majority of the activity in root nodules, and is present almost exclusively in the central tissue of the nodule. However about 20% of NADH-GOGAT I activity is present in the nodule cortex, at about the same specific activity as this isoenzyme is found in the central tissue. Glutamine synthetase (GS; EC 6.3.1.2) occurs predominantly as the γ polypeptide in the central tissue, whereas in the cortex, the enzyme is represented mainly by the β polypeptide. Over 90% of both GS and NADH-GOGAT activities are located in the central tissue of the nodule and GS activity exceeds NADH-GOGAT activity by about twofold in this region. Using the above information, a model for the subcellular location and stoichiometry of nitrogen metabolism in the central tissue of P. vulgaris root nodules is presented.
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  • 34
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    Theoretical and applied genetics 78 (1989), S. 433-435 
    ISSN: 1432-2242
    Keywords: Rhizobium ; Irradiation ; Nitrogen fixation ; Vicia
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Sumary The objective of this work was to know the behaviour and variability of Rhizobium leguminosarum after irradiation. The induced variation was tested under greenhouse conditions on the variety JV 3 of broad beans (Vicia faba) in six replications. Induced genetic variabilty was observed for strain, parent and mutant versus parent. Out of 24 irradiated strains, strain 93-32 performed better with a greater number of nodules and higher dry weight of nodules per plant and biological yield. Environment played an important role in the expression of characters observed. High heritability and genetic advance of these traits indicated that the nitrogen fixation ability of Rhizobium can easily be improved by selection.
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  • 35
    ISSN: 1617-4623
    Keywords: Meiosis ; Saccharomyces cerevisiae ; Sporulation ; Inessential genes ; Meiosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary SPR3 is one of at least nine genes which are expressed in sporulating Saccharomyces cerevisiae cells at the time of meiosis I. We show below that strains homozygous for null alleles of SPR3 are capable of normal meiosis and the production of viable ascospores. We have also monitored SPR3 expression in a series of strains that are defective in meiotic development, using an SPR3: lacZ fusion carried on a single copy plasmid. β-Galactosidase activity occurred at wild-type levels in diploid strains homozygous for mutations in spo13, rad50, rad57 and cdc9, but was greatly reduced in strains carrying cdc8 or spo7 defects. We conclude that SPR3 expression is a valid monitor of early meiotic development, even though the gene is inessential for the sporulation process.
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  • 36
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    Molecular genetics and genomics 217 (1989), S. 149-154 
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; HOM2 gene ; Aspartic semi-aldehyde ; General control
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    Topics: Biology
    Notes: Summary In Saccharomyces cerevisiae the HOM2 gene encodes aspartic semi-aldehyde dehydrogenase (ASA DH). The synthesis of this enzyme had been shown to be derepressed by growth in the presence of high concentrations of methionine. In the present work we have cloned and sequenced the HOM2 gene and found that the promoter region of this gene bears one copy of the consensus sequence for general control of amino acid synthesis. This prompted us to study the regulation of the expression of the HOM2 gene. We have found that ASA DH is the first reported enzyme of the related threonine and methionine pathway to be regulated by the general control of amino acid synthesis.
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  • 37
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    Molecular genetics and genomics 217 (1989), S. 464-470 
    ISSN: 1617-4623
    Keywords: Nucleosome positioning ; LEU2 gene ; Saccharomyces cerevisiae ; tRNA3 Leu gene ; Chromatin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The chromatin structure of theLEU2 gene and its flanks has been studied by means of nuclease digestion, both with micrococcal nuclease and DNase I. The gene is organized in an array of positioned nucleosomes. Within the promoter region, the nucleosome positioning places the regulatory sequences, putative TATA box and upstream activator sequence outside the nucleosomal cores. The tRNA3 Leu gene possesses a characteristic structure and is protected against nucleases. Most of the 5′ flank is sensitive to DNase I digestion, although no clear hypersensitive sites were found. The chromatin structure is independent of either the transcriptional state of the gene or the chromosomal or episomal location. Finally, in the plasmid pJDB207, which lacks most of the promoter, we have found that the chromatin structure of the coding region is similar to that of the wild-type allele.
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  • 38
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    Molecular genetics and genomics 216 (1989), S. 484-491 
    ISSN: 1617-4623
    Keywords: Nitrogen fixation ; nifL ; Repression ; Metal ions
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The ability of the Klebsiella pneumoniae nifL gene product to antagonise NIFA mediated transcriptional activation from the nifH promoter in vivo was inhibited either by metal deprivation, or by the presence of the iron chelators EDDA or Desferal in the growth medium. This inhibition of the repressive activity of NIFL was reversed by the addition of ferrous or manganous ions to the medium but was unaffected by other transition metals. The dependence on metal ions for NIFL activity was observed when NIFL was overexpressed and when cultures were exposed to oxygen or high levels of fixed nitrogen. Immunochemical evidence suggests that NIFL and NIFA associate to form a functional protein complex. Metal ions are apparently not required for the formation of this complex.
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  • 39
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    Molecular genetics and genomics 216 (1989), S. 511-516 
    ISSN: 1617-4623
    Keywords: Nonsense mutation ; Read-through ; Saccharomyces cerevisiae
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    Topics: Biology
    Notes: Summary The yeast invertase structural gene SUC2 has two naturally occurring alleles, the active one and a silent allele called suc2°. Strains carrying suc2° are unable to ferment sucrose and do not show detectable invertase activity. We have isolated suc2° and found an amber codon at position 232 of 532 amino acids. However, transformants carrying suc2° on a multicopy plasmid were able to ferment sucrose and showed detectable invertase activity. Full-length invertase was found in gels stained for active invertase and in immunoblots. Therefore we concluded that the amber codon is occasionally read as an amino acid. The calculated frequency of read-through is about 4% of all translation events.
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  • 40
    ISSN: 1617-4623
    Keywords: glnB ; Klebsiella pneumoniae ; Nitrogen control ; Glutamine synthetase ; Nitrogen fixation
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    Notes: Summary The role of theKlebsiella pneumoniae PII protein (encoded byglnB) in nitrogen regulation has been studied using two classes ofglnB mutants. In Class I mutants PII appears not to be uridylylated in nitrogen-limiting conditions and in Class II mutants PII is not synthesised. The effects of these mutations on expression from nitrogen-regulated promoters indicate that PII is not absolutely required for nitrogen control. Furthermore the uridylylated form of PII(PII-UMP) plays a significant role in the response to changes in nitrogen status by counteracting the effect of PII on NtrB-mediated dephosphorylation of NtrC. PII is not involved in thenif-specific response to changes in nitrogen status mediated by NifL.
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  • 41
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    Molecular genetics and genomics 218 (1989), S. 293-301 
    ISSN: 1617-4623
    Keywords: Sporulation ; DNA sequence ; Saccharomyces cerevisiae ; Meiosis ; Chromosome segregation
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    Topics: Biology
    Notes: Summary Genetic studies have previously demonstrated that the RED1 gene of Saccharomyces cerevisiae is required for chromosome segregation at the first meiotic division. Northern blot hybridization analysis indicates that the RED1 gene produces two transcripts of 2.75 and 3.2 kilobases. The major 2.75 kb transcript is not present in mitotic cells and is meiotically induced to accumulate maximally just prior to the meiosis I division. The DNA sequence of the RED1 gene was determined and used to predict the amino acid sequence of the encoded gene product. The RED1 protein is 827 amino acids in length and has a molecular weight of 95.5 kilodaltons. There is no significant homology between the RED1 amino acid sequence and other known protein sequences, including those encoded by genes essential for meiosis.
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  • 42
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    Molecular genetics and genomics 215 (1989), S. 401-406 
    ISSN: 1617-4623
    Keywords: Protein secretion ; Saccharomyces cerevisiae ; Invertase ; Endoplasmic reticulum ; Secretion mutants
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    Topics: Biology
    Notes: Summary Intercompartmental transport of secreted proteins in yeast was analysed using invertase mutants. Deletions and insertions at the BamHI (position +787) or the Asp718 (position +1159) sites of the SUC2 gene led to mutant proteins with different behaviour regarding secretion, localization and enzyme activity. The deletion mutants showed accumulation of core glycosylated material in the endoplasmic reticulum (ER) a decrease of secreted protein by 5%–30% and loss of enzyme activity. The secreted material was localized in the culture medium and not — as is normal for invertase-in the cell wall. No delay in transport from the Golgi to the cell surface was observed, indicating that the rate-limiting step for secretion is at the ER-Golgi stage. Two insertion mutants, pIPA and pIPB, retained enzyme activity. Mutant pIPB showed 10% secretion, while 60%–70% secretion was observed for pIPA. While the non-secreted material accumulated in the ER, the secreted material was present in the cell wall. The results suggest that the presence of structures incompatible with secretion leads to ER accumulation of mutated invertase.
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  • 43
    ISSN: 1617-4623
    Keywords: RNA splicing ; Maturase ; Recombinase ; Mitochondria ; Saccharomyces cerevisiae
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    Notes: Summary When the bI4 RNA maturase, encoded by the fourth intron of the mitochondrial cytochrome b gene of Saccharomyces cerevisiae, was expressed in Escherichia coli, formation of intra-chromosomal Lac+ recombinants was stimulated threefold. This “hyper-rec” phenotype was recA as well as recBCD dependent. The most active form of the bI4 maturase stimulated homologous recombination whereas splicing deficient mutants of bI4 maturase were either deficient in or unable to stimulate homologous recombination.
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  • 44
    ISSN: 1617-4623
    Keywords: Nucleotide sequence ; PET gene ; Mitochondrial import ; Saccharomyces cerevisiae
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    Notes: Summary Translation of mitochondrial cytochrome b mRNA in yeast is activated by the product of the nuclear gene CBS1. CBS1 encodes a 27 kDa precursor protein, which is cleaved to a 24 kDa mature protein during the import into isolated mitochondria. The sequences required for mitochondrial import reside in the amino-terminal end of the CBS1 precursor. Deletion of the 76 amino-terminal amino acids renders the protein incompetent for mitochondrial import in vitro and non-functional in vivo. When present on a high copy number plasmid and under the control of a strong yeast promoter, biological function can be restored by this truncated derivative. This observation indicates that the CBS1 protein devoid of mitochondrial targeting sequences can enter mitochondria in vivo, possibly due to a bypass of the mitochondrial import system.
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  • 45
    ISSN: 1617-4623
    Keywords: Rhodobacter capsulatus ; Nitrogen fixation ; DNA sequence analysis ; nifE, nifN, nifX genes ; Protein comparisons
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary Rhodobacter capsulatus genes homologous to Klebsiella pneumoniae nifE, nifN and nifX were identified by DNA sequence analysis of a 4282 bp fragment of nif region A. Four open reading frames coding for a 51188 (NifE), a 49459 (NifN), a 17459 (NifX) and a 17472 (ORF4) dalton protein were detected. A typical NifA activated consensus promoter and two imperfect putative NifA binding sites were located in the 377 bp sequence in front of the nifE coding region. Comparison of the deduced amino acid sequences of R. capsulatus NifE and NifN revealed homologies not only to analogous gene products of other organisms but also to the α and β subunits of the nitrogenase iron-molybdenum protein. In addition, the R. capsulatus nifE and nifN proteins shared considerable homology with each other. The map position of nifX downstream of nifEN corresponded in R. capsulatus and K. pneumoniae and the deduced molecular weights of both proteins were nearly identical. Nevertheless, R. capsulatus NifX was more related to the C-terminal end of NifY from K. pneumoniae than to NifX. A small domain of approximately 33 amino acid residues showing the highest degree of homology between NifY and NifX was also present in all nifB proteins analyzed so far. This homology indicated an evolutionary relationship of nifX, nifY and nifB and also suggested that NifX and NifY might play a role in maturation and/or stability of the iron-molybdenum cofactor. The open reading rame (ORF4) downstream of nifX in R. capsulatus is also present in Azotobacter vinelandii but not in K. pneumoniae. Interposon-induced insertion and deletion mutants proved that nifE and nifN were necessary for nitrogen fixation in R. capsulatus. In contrast, no essential role could be demonstrated for nifX and ORF4 whereas at least one gene downstream of ORF4 appeared to be important for nitrogen fixation.
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  • 46
    ISSN: 1617-4623
    Keywords: Mitochondrial introns ; Reverse transcriptase ; Saccharomyces cerevisiae
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    Notes: Summary Some pet- (or mit-) mutations impeding the splicing of one or several intron(s) of the yeast mitochondrial pre-mRNA(s) are suppressed in vivo by the DNA deletion of these introns. We have genetically demonstrated that introns aI1 and/or aI2 of the cytochrome c oxidase subunit 1 gene are necessary for this deletion process. The facts that adjacent introns are simultaneously deleted and that, in the pet- (or mit-) mutants which easily revert by intron deletion, the splicing of the introns they affect is only partially blocked, suggest that the intron encoded proteins aI1 and/or aI2 could intervene by means of their putative reverse transcriptase activity.
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  • 47
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    Protoplasma 150 (1989), S. 19-26 
    ISSN: 1615-6102
    Keywords: Nitrogen fixation ; Peanut ; Root nodules ; Dense body ; Microbody ; Oleosome
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    Topics: Biology
    Notes: Summary Nitrogen-fixing peanut root nodules are characterized by their unique structural organization, distinct from other legume nodules. The focus of this study has been in and around the hostsymbiont interface, where the bacterioid and the host cell surface (peribacteroid membrane envelope) interact during symbiosis. The infected nodule cells have revealed the presence of lipid bodies (oleosomes) in intimate association with the peribacteroid membrane, which encloses the large spherical bacteroids with a relatively narrow peribacteroid space. Electron dense structures, referred to as dense bodies have been found attached to the bacteroid outer membranes at the host-symbiont interface. The dense bodies are osmiophilic, amorphous and 3,3′-diaminobenzidine positive. The isolated intact bacteroids with dense bodies attached to their cell wall showed significant catalase activity. Many microbodies showing DAB-positive reaction have been found in the host cytoplasm, associated closely with the peribacteroid membrane. These ultrastructural and cytochemical characteristics of peanut root nodules suggest that lipids are utilized during symbiosis and the dense bodies and microbodies may be involved in the catabolic process.
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  • 48
    ISSN: 1615-6102
    Keywords: Microtubule neoformation ; Nocodazole ; Protoplasts ; Saccharomyces cerevisiae
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    Notes: Summary By following microtubule neoformation after their complete destruction by nocodazole, we analyzed the pattern of microtubule nucleation in protoplasts ofSaccharomyces cerevisiae. Using immunofluorescence, the drug was shown to induce rapid and complete disassembly of both cytoplasmic and spindle microtubules and to selectively block protoplast nuclear division at a defined stage of the cell cycle. Treated protoplasts placed in a drug-free environment recovered a more abundant microtubular system. The majority of microtubules re-formed at SPBs whereas a minority of free-ended microtubules nucleated in the cytoplasm of the protoplasts without any detectable association with recognizable nucleation sites. Random nucleation of free microtubules might be induced by high amounts of unpolymerized tubulin likely to be present in the protoplasts at the moment of drug release.
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  • 49
    ISSN: 1615-6102
    Keywords: Plectonema boryanum ; Cyanobacteria ; Ultrastructure ; Nitrogen fixation ; Nitrogen starvation ; Immunogold localization
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    Notes: Summary The ultrastructure of fructose-supplemented and unsupplemented nitrogen-fixing (fix +) and nonfixing (fix −)Plectonema boryanum UTEX 581 cells was examined by transmission electron microscopy. The most prominent structural differences included the arrangement and morphology of the thylakoids and alterations in the appearance of the interthylakoidal spaces. These ultrastructural differences, together with other observations such as glycogen content and presence of nitrogenase (using acetylene reduction assay and immunogold localization), readily distinguished nonfixingP. boryanum from nitrogen-fixing cells.
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  • 50
    ISSN: 1615-6102
    Keywords: Legumes ; Nitrogen fixation ; Nodulation ; Rhizobium ; Symbiosis
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    Notes: Summary Two methods have been developed in order to discriminate between lateral roots, nodules and root-derived structures which exhibit both root and nodule histological features and which can develop on legumes inoculated with certainRhizobium mutants. The first method, known as the “clearing method”, allows the observation by light microscopy of cleared undissected root-structures. The second, known as the “slicing method”, is a complementary technique which provides a greater degree of structural information concerning such structures. The two methods have proved invaluable in defining unequivocally the nature of the interaction between a rhizobial strain and a legume host.
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  • 51
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    Molecular genetics and genomics 218 (1989), S. 531-535 
    ISSN: 1617-4623
    Keywords: Origin of replication ; Promoter ; Saccharomyces cerevisiae
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    Notes: Summary In an attempt to devise a new assay for ARS-binding proteins we have inserted the HO ARS between the upstream activation site and the TATA region of the yeast CYC1 promoter. A marked reduction in promoter activity is observed. Inactivation of the HO ARS element by point mutation does not restore promoter activity to its original level, although a modest activation is seen. We have also inserted the HO ARS into the intron of the yeast actin gene; although there is no apparent deleterious effect on transcription, the activity of the ARS is abolished in this new environment.
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  • 52
    ISSN: 1617-4623
    Keywords: Rhizobium leguminosarum ; Nitrogen fixation ; nif/fix genes ; Escherichia coli minicells ; Transcription regulation
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    Notes: Summary On the Rhizobium leguminosarum PRE sym plasmid, fixABC and a novel gene fixW were identified upstream of the regulatory gene nifA. The molecular masses of FixABC, 29, 44 and 50 kDa respectively, were estimated by polyacrylamide gel electrophoresis (PAGE) and of FixW, 25 kDa, by PAGE and nucleotide sequencing. Hybridization studies using bacteroid mRNA as a probe showed that fixABC is one operon which can be transcribed independently of fixW. Nucleotide sequencing revealed that both fixW and fixA are preceded by a nif consensus promoter. The fixA promoter partly overlaps the 3′-terminal coding region of fixW, indicating that readthrough from fixW into fixA is possible. Two open reading frames, ORF71 and ORF79, precede fixW and form one operon with fixW. ORF71 contains sequences homologous to the fixA promoter and 5′-terminal coding region. One more duplication of fixA sequences was detected, also located within the sym plasmid nif/fix clusters. One duplication of fixW sequences was found. No fixW homologue could be found in other nitrogen fixing organisms except in a number of R. leguminosarum strains.
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  • 53
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Heavy metal resistance ; DNA sequence ; Membrane
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    Topics: Biology
    Notes: Summary A DNA fragment conferring resistance to zinc and cadmium ions in the yeast Saccharomyces cerevisiae was isolated from a library of yeast genomic DNA. Its nucleotide sequence revealed the presence of a single open reading frame (ORF; 1326 bp) having the potential to encode a protein of 442 amino acid residues (molecular mass of 48.3 kDa). A frameshift mutation introduced within the ORF abolished resistance to heavy metal ions, indicating the ORF is required for resistance. Therefore, we termed it the ZRC1 (zinc resistance conferring) gene. The deduced amino acid sequence of the gene product predicts a rather hydrophobic protein with six possible membrane-spanning regions. While multiple copies of the ZRC1 gene enable yeast cells to grow in the presence of 40 mM Zn2+, a level at which wild-type cells cannot survive, the disruption of the chromosomal ZRC1 locus, though not a lethal event, makes cells more sensitive to zinc ions than are wild-type cells.
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  • 54
    ISSN: 1617-4623
    Keywords: Nitrogen fixation ; draT ; draG ; TTG initiation codon
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    Topics: Biology
    Notes: Summary Nitrogen fixation activity in the photosynthetic bacterium Rhodospirillum rubrum is controlled by the reversible ADP-ribosylation of the dinitrogenase reductase component of the nitrogenase enzyme complex. This report describes the cloning and characterization of the genes encoding the ADP-ribosyltransferase (draT) and the ADP-ribosylglycohydrolase (draG) involved in this regulation. These genes are shown to be contiguous on the R. rubrum chromosome and highly linked to the nifHDK genes. Sequence analysis revealed the use of TTG as the initiation codon of the draT gene as well as a potential open reading frame immediately downstream of draG. The mono-ADP-ribosylation system in R. rubrum is the first in which both the target protein and modifying enzymes as well as their structural genes have been isolated, making it the model system of choice for analysis of this post-translational regulatory mechanism.
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  • 55
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; a-factor ; Conjugation ; G1 arrest ; ssl mutations
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Nine independent mutants which are supersensitive (ssl −) to G1 arrest by the mating hormone a-factor were isolated by screening mutagenized Saccharomyces cerevisiae MATα cells on solid medium for increased growth inhibition with a-factor. These mutants carried lesions in two complementation groups, ssl1 and ssl2. Mutations at the ssl1 locus were mating type specific: MATα ssl1 − cells were supersensitive to α-factor but MATα ssl1 − were not supersensitive to α-factor. In contrast, mutations at the ssl2. locus conferred supersensitivity to the mating hormone of the opposite mating type on both MATα, and MATa cells. The α-cell specific capacity to inactivate externally added a-factor was shown to be lacking in MATα ssl1 − mutants whereas MATα ssl2. cells were able to inactivate a-factor. Complementation analysis showed that ssl2 and sst2, a mutation originally isolated as conferring supersensitivity to α-factor to MATa cells, are lesions in the same gene. The ssl1 gene was mapped 30.5 centi-Morgans distal to ilv5 on chromosome XII.
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  • 56
    ISSN: 1617-4623
    Keywords: Aspergillus oryzae ; Alkaline protease ; Prepro sequence ; Heterologous expression ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have cloned and determined the nucleotide sequence of a cDNA fragment for the entire coding region of the alkaline protease (Alp) from a filamentous ascomycete Aspergillus oryzae. According to the deduced amino acid sequence, Alp has a putative prepro region of 121 amino acids preceding the mature region, which consists of 282 amino acids. A consensus sequence of a signal peptide consiting of 21 amino acids is found at the N-terminus of the prepro region. The primary structure of the mature region shares extensive homology (29%–44%) with those of subtilisin families, and the three residues (Asp 32, His 64 and Ser 221 in subtilisin BPN′) composing the active site are preserved. The entire cDNA, coding for prepro Alp, when introduced into the yeast Saccharomyces cerevisiae, directed the secretion of enzymatically active Alp into the culture medium, with its N-terminus and specific activity identical to native Aspergillus Alp.
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  • 57
    ISSN: 1617-4623
    Keywords: Gene conversion ; Crossing-over ; Mismatch repair ; Saccharomyces cerevisiae ; Psoralens
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The induction of gene conversion and mitotic crossing-over by photoaddition of psoralens, 254 nm ultraviolet radiation, and nitrogen mustards was determined in diploid cells homozygous for the pso3-1 mutation and in the corresponding wild type of Saccharomyces cerevisiae. For these different agents, the frequency of non-reciprocal events (conversion) is reduced in the pso3-1 mutant compared to the wild type. In contrast, the frequency of reciprocal events (crossing-over) is increased at a range of doses. These observations, together with the block in induced mutagenesis for both reverse and forward mutations previously reported for the pso3-1 mutant, suggest that the PS03 gene product plays a role in mismatch repair of short patch regions. The block in gene conversion in the pso3 homozygous diploid leads, in the case of nitrogen mustards, to specific repair intermediates which are lethal to the cells.
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  • 58
    ISSN: 1617-4623
    Keywords: Enhanced secretion ; Human lysozyme production ; Protease mutant ; Protein processing ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Yeast mutant strains which secrete large amounts of human lysozyme were screened using an agar medium containing bacterial cells. Nine mutants secreted over 10 times more lysozyme than the wild-type parent strain. The mRNA levels for lysozyme in the mutants were not higher than that of the wild-type strain. Three of the mutant strains were deficient in carboxypeptidase Y activity. It was found that the protease deficiency was caused by a deficiency in conversion of proenzyme to mature enzyme in ssl1 mutant cells. The ssl1 gene was found to be closely linked to the centromere and determine both the efficiency of secretion of lysozyme and the processing of carboxypeptidase Y.
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  • 59
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    Molecular genetics and genomics 219 (1989), S. 495-498 
    ISSN: 1617-4623
    Keywords: ARS1 ; Plasmid multimerization ; RAD52 ; Saccharomyces cerevisiae ; Eukaryotic recombination
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A mutant plasmid, pX, derived from the 1453 base pair small plasmid, YARp1 (or TRP1 RI circle), consists of 849 base pairs of DNA bearing the TRP1 gene and the ARS1 sequence of Saccharomyces cerevisiae and, unlike YARp1 and other commonly used yeast plasmids, highly multimerizes in a S. cerevisiae host. The multimerization of pX was dependent on RAD52, which is known to be necessary for homologous recombination in S. cerevisiae. Based upon this observation, a regulated system of multimerization of pX with GAL1 promoter-driven RAD52 has been developed. We conclude that the regulated multimerization of pX could provide a useful model system to study genetic recombination in the eukaryotic cell, in particular to investigate recombination intermediates and the effects of various trans-acting mutations on the multimerization and recombination of plasmids.
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  • 60
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    Molecular genetics and genomics 215 (1989), S. 455-462 
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Orotate phosphoribosyl transferase ; Nucleotide sequence ; Transcription ; Regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The URA5 gene of Saccharomyces cerevisiae encodes orotate phosphoribosyl transferase (EC 2.4.2.10; OPRTase) which catalyses the transformation of orotate to OMP in the pyrimidine pathway. We present in this paper the cloning and the sequencing of this gene, the last in the yeast pyrimidine pathway to be cloned. We have deduced the protein sequence of the OPRTase of S. cerevisiae from the DNA sequence and compared it to that of Escherichia coli, Podospora anserina and Dictyostelium discoideum. Some important similarities in the structure of these four proteins have been found. Finally, we have quantified the transcription of the URA5 gene in different physiological conditions and confirmed that it was not under the control of UTP or any intermediary product of the pathway.
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  • 61
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    Molecular genetics and genomics 215 (1989), S. 507-516 
    ISSN: 1617-4623
    Keywords: Nitrogen fixation ; Regulation ; Rhodobacter capsulatus ; Gene sequences ; Transcription factors
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have determined the DNA sequence for the genes nifR1, nifR2 and nifR4 in the photosynthetic bacterium Rhodobacter capsulatus. These genes regulate transcription of the nifHDK operon and so limit the expression of nitrogen fixation activity to periods of low environmental concentrations of both oxygen and fixed nitrogen. The sequences of these three genes are similar to components of the ntr regulation system in Escherichia coli and Klebsiella pneumoniae. The two-component regulatory system of ntrB and ntrC in E. coli is represented by nifR2 and nifR1 in R. capsulatus and nifR4 in R. capsulatus is the equivalent of the E. coli ntr-related sigma factor ntrA.
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  • 62
    ISSN: 1617-4623
    Keywords: DNA sequence ; PET gene ; Saccharomyces cerevisiae ; Mitochondrial import ; cytochrome c oxidase
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    Notes: Summary The yeast nuclear SCO1 gene is required for accumulation of the mitochondrially synthesized cytochrome c oxidase subunits I and II (COXI and COXII). We cloned and characterized the SCO1 gene. It codes for a 0.9 kb transcript. DNA sequence analysis predicts a 33 kDa protein. As shown by in vitro transcription and translation experiments in combination with import studies on isolated mitochodria, this protein is matured into a 30 kDa polypeptide which is tightly associated with a mitochondrial membrane. The possible function of the SCO1 gene product in the assembly of cytochrome c oxidase is discussed.
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  • 63
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; GLN1 ; Glutamine synthetase ; Regulatory systems ; Transcription
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary TheGLN1 gene ofSaccharomyces cerivisiae was cloned by complementation of agln1 auxotroph. AGLN1-lacZ fusion was constructed to assayGLN1 promoter activity. β-Galactosidase and glutamine synthetase expression in chromosomally integratedGLN1-lacZ fusion strains were co-regulated in response to a shift from glutamine to glutamate as the nitrogen source, purine limitation, and 3-aminotriazole-induced histidine starvation. Regulation ofGLN1 expression by each of the three pathways occurred at the transcriptional level. Increased accumulation ofGLN1 mRNA was observed within 5 min after a shift from glutamine to glutamate as the nitrogen source. After 5 min following glutamine addition to the cells growing with glutamate as nitrogen source. This indicates that theGLN1 message is unstable and has a half-life of approximately 3 min. Deletion analysis indicated that the sequences required forGLN1 expression are located within approximately 350 bp upstream from the transcriptional initiation site.
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  • 64
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; DNA repair ; Cross-link ; Transposon mapping ; Nitrogen mustard
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have isolated yeast gene SNM1 via complementation of sensitivity towards bi- and tri-functional alkylating agents in haploid and diploid yeast DNA repair-deficient snm1-1 mutants. Four independent clones of plasmid DNA containing the SNM1 locus were isolated after transformation with a YEp24-based yeast gene bank. Subcloned SNM1-containing DNA showed (i) complementation of the repair-deficiency phenotype caused by either one of the two different mutant alleles snm1-1 and snm1-2 ts; (ii) complementation in haploid and diploid yeast snm1-1 mutants by either single or multiple copies of the SNM1 locus; and (iii) that the SNM1 gene is at most 2.4 kb in size. Expression of SNM1 on the smallest subclone, however, was under the control of the GAL1 promotor. Gene size and direction of transcription was further verified by mutagenesis of SNM1 by Tn10-LUK transposon insertion. Five plasmids containing Tn10-LUK insertions at different sites of the SNM1-containing DNA were able to disrupt function of genomic SNM1 after gene transplacement. Correct integration of the disrupted SNM1::Tn10-LUK at the genomic site of SNM1 was verified via tetrad analysis of the sporulated diploid obtained after mating of the SNM1::Tn10-LUK transformant to a haploid strain containing the URA3 SNM1 wild-type alleles. The size of the poly(A)+ RNA transcript of the SNM1 gene is 1.1 kb as determined by Northern analysis.
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  • 65
    ISSN: 1617-4623
    Keywords: pso4-1 mutation ; Saccharomyces cerevisiae ; Error-prone recombinational repair ; Mitotic recombination
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The induction of mitotic gene conversion and crossing-over inSaccharomyces cerevisiae diploid cells homozygous for thepso4-1 mutation was examined in comparison to the corresponding wild-type strain. Thepso4-1 mutant strain was found to be completely blocked in mitotic recombination induced by photoaddition of mono- and bifunctional psoralen derivatives as well as by mono- (HN1) and bifunctional (HN2) nitrogen mustards or 254 nm UV radiation in both stationary and exponential phases of growth. Concerning the lethal effect, diploids homozygous for thepso4-1 mutation are more sensitive to all agents tested in any growth phase. However, this effect is more pronounced in the G2 phase of the cell cycle. These results imply that the ploidy effect and the resistance of budding cells are under the control of thePSO4 gene. On the other hand, thepso4-1 mutant is mutationally defective for all agents used. Therefore, thepso4-1 mutant has a generalized block in both recombination and mutation ability. This indicates that thePSO4 gene is involved in an error-prone repair pathway which relies on a recombinational mechanism, strongly suggesting an analogy between thepso4-1 mutation and theRecA orLexA mutation ofEscherichia coli.
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  • 66
    ISSN: 1617-4623
    Keywords: GC clusters ; Mobile elements ; Target sites ; mtDNA ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary GC clusters constitute the major repetitive elements in the mitochondrial (mt) genome of the yeast Saccharomyces cerevisiae. Many of these clusters are optional and thus contribute much to the polymorphism of yeast mtDNAs. We have made a systematic search for polymorphic sites by comparing mtDNA sequences of various yeast strains. Most of the 26 di- or polymorphic sites found differ by the presence or absence of a GC cluster of the majority class, here referred to as the M class, which terminate with an AGGAG motif. Comparison of sequences with and without the GC clusters reveal that elements of the subclasses M1 and M2 are inserted 3′ to a TAG, flanked by A+T rich sequences. M3 elements, in contrast, only occur in tandem arrays of two to four GC clusters; they are consistently inserted 3′ to the AGGAG terminal sequence of a preexisting cluster. The TAG or the terminal AGGAG, therefore, are regarded as being part of the target sites for M1 and M2 or M3 elements, respectively. The dinucleotide AG is in common to both target sites; it also occurs at the 3′ terminus (AGGAG). This suggests its duplication during GC cluster insertion. This notion is supported by the observation that GC clusters of the minor classes G and V similarily repeat at their 3′ terminus a GT or an AA dinucleotide, respectively, from their putative target sites.
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  • 67
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    Molecular genetics and genomics 218 (1989), S. 57-63 
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; PET gene ; Transcriptional regulation ; Anaerobiosis ; 5′ mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Translation of mitochondrial cytochrome b RNA in yeast requires the product of the nuclear gene CBS1, a 27.5 kDa soluble mitochondrial protein. In this paper we show that the CBS1 gene is located on chromosome IV immediately adjacent to COX9, the gene coding for cytochrome c oxidase subunit VIIa. CBS1 is transcribed as a very low abundant 900 b RNA. Transcription starts at a single position 101 bp upstream of the CBS1 initiation codon. At positions-39 to-27 of its leader sequence it contains a small open reading frame of 4 codons. By monitoring the β-galactosidase activity of a CBS1/lacZ fusion construct we show that expression of CBS1 is subjected to regulation by oxygen and by glucose: the β-galactosidase activity is elevated threefold in glycerol or galactose grown cells compared to that in glucose grown cells. A further threefold reduction of the activity is observed in anaerobically grown cells. In accordance with this result is the observation that the steady-state level of CBS1 mRNA of anaerobically grown cells is ninefold lower than that of aerobically cultured cells.
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  • 68
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    Plant and soil 114 (1989), S. 63-68 
    ISSN: 1573-5036
    Keywords: Azolla pinnata ; Nitrogen fixation ; N yield ; Oryza sativa ; Urea-N
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Application of 0, 30, 60, 90 and 120 kg N ha−1 of urea (U) in split doses with (and without)Azolla pinnata, R. Brown was studied for three consecutive seasons under planted field condition. Fresh weight (FW), acetylene reduction activity (ARA) and N yield of Azolla were found to be maximum 14 days after inoculation (DAI). Among the different treatments, maximum Azolla growth was recorded in no N control. The FW, ARA and N yield of Azolla were inhibited increasingly with the increase in N levels. Irrespective of season, FW and N yield of Azolla were inhibited only a small extent with 90 kg N ha−1 U, beyond which the inhibition was pronounced. ARA was inhibited only slightly up to 60 kg N ha−1 of U. Grain yield and crop N uptake of rice increased significantly up to 90 kg N ha−1 of U (alone or in combination with Azolla) in the dry seasons (variety IR 36) and up to 60 kg N ha−1 U in the wet season (variety CR 1018).
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  • 69
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    Journal of bioenergetics and biomembranes 21 (1989), S. 589-603 
    ISSN: 1573-6881
    Keywords: Vacuolar membrane H+ATPase ; vacuoles ; Saccharomyces cerevisiae ; catalytic cooperativity of ATP hydrolysis ; VMA genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Our current work on a vacuolar membrane proton ATPase in the yeastSaccharomyces cerevisiae has revealed that it is a third type of H+-translocating ATPase in the organism. A three-subunit ATPase, which has been purified to near homogeneity from vacuolar membrane vesicles, shares with the native, membrane-bound enzyme common enzymological properties of substrate specificities and inhibitor sensitivities and are clearly distinct from two established types of proton ATPase, the mitochondrial F0F1-type ATP synthase and the plasma membrane E1E2-type H+-ATPase. The vacuolar membrane H+-ATPase is composed of three major subunits, subunita (M r =67 kDa),b (57kDa), andc (20 kDa). Subunita is the catalytic site and subunitc functions as a channel for proton translocation in the enzyme complex. The function of subunitb has not yet been identified. The functional molecular masses of the H+-ATPase under two kinetic conditions have been determined to be 0.9–1.1×105 daltons for single-cycle hydrolysis of ATP and 4.1–5.3×105 daltons for multicycle hydrolysis of ATP, respectively.N,N′-Dicyclohexylcarbodiimide does not inhibit the former reaction but strongly inhibits the latter reaction. The kinetics of single-cycle hydrolysis of ATP indicates the formation of an enzyme-ATP complex and subsequent hydrolysis of the bound ATP to ADP and Pi at a 7-chloro-4-nitrobenzo-2-oxa-1,3-diazolesensitive catalytic site. Cloning of structural genes for the three subunits of the H+-ATPase (VMA1, VMA2, andVMA3) and their nucleotide sequence determination have been accomplished, which provide greater advantages for molecular biological studies on the structure-function relationship and biogenesis of the enzyme complex. Bioenergetic aspects of the vacuole as a main, acidic compartment ensuring ionic homeostasis in the cytosol have been described.
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  • 70
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    Journal of bioenergetics and biomembranes 21 (1989), S. 621-632 
    ISSN: 1573-6881
    Keywords: ATPase ; [H+]-ATPase ; proton transport ; Neurospora crassa ; Saccharomyces cerevisiae ; Schizosaccharomyces pombe
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract The fungal plasma membrane contains a proton-translocating ATPase that is closely related, both structurally and functionally, to the [Na+, K+]-, [H+, K+]-, and [Ca2+]-ATPases of animal cells, the plasma-membrane [H+]-ATPase of higher plants, and several bacterial cation-transporting ATPases. This review summarizes currently available information on the molecular genetics, protein structure, and reaction cycle of the fungal enzyme. Recent efforts to dissect structure-function relationships are also discussed.
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  • 71
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    Biology and fertility of soils 6 (1988), S. 39-44 
    ISSN: 1432-0789
    Keywords: Alnus ; Energy forestry ; Frankia ; Meadow soil ; Nitrogen fixation ; Nodulation ; Peat soil
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Use of the N2-fixing grey alder, Alnus incana (L.) Moench, as a short-rotation crop for energy production is currently being explored. To evaluate the need for inoculation of alders, the distribution of infective propagules of Frankia in the soil at potential sites for alder plantations was examined. Uninoculated grey alder seedlings were grown in three types of soil. Frequent nodulation was found in a meadow soil which had been free from actinorhizal plants for nearly 60 years, but the alder seedlings failed to nodulate in peat soil from two different bog sites. One of these bogs had been exploited for peat and the surface layer of the peat had been removed, so that the soil samples were taken from deep layers of the peat. At the other site, an area of cultivated peat, there were no infective propagules of Frankia in plots without alders; the infective Frankia was present in plots only where it had been introduced by inoculated alders. There was no detectable air-borne dispersal of Frankia. Instead, water movement might account for the dispersal of Frankia in peat. Although the apparent absence of Frankia in these peat soils necessitates inoculation of alder seedlings before planting out, this makes it possible to introduce and maintain Frankia strains with selected beneficial characteristics, since there is no competition from an indigenous Frankia flora.
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  • 72
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    Biology and fertility of soils 6 (1988), S. 279-281 
    ISSN: 1432-0789
    Keywords: Sesbania rostrata ; Green manure ; Biofertilizer ; Nitrogen fixation ; Stem nodule
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Ratooning and stem cutting were compared with seeding in order to reduce the amount of seeds of Sesbania rostrata for green-manure growth. Both methods increased the biofertilizer yield highly significantly within a 6-week growth period.
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  • 73
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Site specific recombination ; 2 μ DNA plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The purpose of this work is to identify and quantitate in vivo 2 μ plasmid FLP-independent recombination in yeast, using a nonselective assay system for rapid detection of phenotypic expression of the recombination events. A tester plasmid was constructed such that in vivo recombination between 2 μ direct repeat sequences produces the resolution of the plasmid into two circular DNA molecules. This recombinational event is detected as a phenotypic shift from red to white colonies, due to the mitotic loss of the plasmid portion containing the yeast ADE8 gene in a recipient ade1 ade2 ade8 genetic background. In the absence of the 2 μ FLP recombinase and/or its target DNA sequence, recombination is not abolished but rather continues at a high frequency of about 17%. This suggests that the FLP-independent events are mediated by the chromosomally-encoded general homologous recombination system. We therefore conclude that the totality of 2 μ DNA recombination events occurring in FLP+ cells is the contribution of both FLP-mediated and FLP-independent events.
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  • 74
    ISSN: 1432-0983
    Keywords: Schizosaccharomyces pombe ; Saccharomyces cerevisiae ; Argininosuccinate lyase ; Gene cloning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A gene bank of Sau3A partially restricted Schizosaccharomyces pombe DNA in YEp13 was used to transform an arg4 mutant of Saccharomyces cerevisiae. One colony was recovered which contained the YEp13 plasmid bearing a large insert complementing the argininosuccinate lyase (ASL) mutation. As shown by restriction mapping and subcloning experiments, the DNA sequence required for complementation is localized on a 2 kb BamHI-BamHI fragment. The plasmid complemented several S. cerevisiae arg4 mutants of independent origin and a S. pombe arg7 mutant lacking ASL. Low but significant ASL activities were detected in crude extracts of these transformants. No complementation of the E. coli argH mutant was observed. Southern blot hybridizations showed that the insert originates from the S. pombe genome. No cross-hybridization was found between this sequence and S. cerevisiae DNA. It can be concluded that the cloned DNA fragment bears the S. pombe ARG7 gene coding for ASL.
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  • 75
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    Current genetics 14 (1988), S. 413-418 
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Chromosome length polymorphisms ; FIGE ; OFAGE
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Although Saccharomyces cerevisiae strains generally have similar chromosomal band patterns as revealed by pulsed field gel electrophoresis, individual bands often move slightly differently from one strain to the other. Surveying strains from our stock collection, we found that nearly all the bands of a certain pair of strains differed in their mobility. Some of these chromosome length polymorphisms segregated in a 2:2 ratio, indicating that they resulted from single structural alterations (i.e. additions or deletions). One of these was mapped on the right arm of chromosome 1. Others did not segrate in a simple 2:2 ratio. That is, there were progenies which had bands not present in either parent. We suggest that these new bands are the products of recombination between homologous chromosomes having two or more structural alterations.
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  • 76
    ISSN: 1432-0983
    Keywords: ARS ; Saccharomyces cerevisiae ; Schizosaccharomyces pombe ; Tetrahymena thermophila
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have isolated several Tetrahymena thermophila chromosomal DNA fragments which function as autonomously replicating sequences (ARS) in the heterologous Saccharomyces cerevisiae and Schizosaccharomyces pombe selection systems. The Tetrahymena ARS sequences were first isolated in S. cerevisiae and were derived from non-ribosomal micro- and macronuclear DNA. Sequence analysis of the ARS elements identified either perfect or close matches with the 11 by S. cerevisiae ARS core consensus sequence. Subcloning studies of two Tetrahymena ARS elements defined functional regions ranging in size from 50 to 300 bp. Testing of the ARS elements in S. pombe revealed that most of the T. thermophila inserts confer ARS function in both yeasts, at least in the sense of promoting a high transformation frequency to plasmids which contain them. However, the actual sequences responsible for ARS activity were not always identical in the two yeasts.
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  • 77
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Ribosomal protein genes ; CYH2
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A diploid strain of the yeast Saccharomyces cerevisiae has been constructed that has one copy of the ribosomal protein gene CYH2 completely deleted and replaced with the TRP1 gene using the method of Rothstein (1983). There are only small differences in growth rate and no detectable difference in steady state level of CYH2 mRNA between the diploid that is heterozygous for the CYH2 deletion and the parent diploid with two normal copies of this gene. This suggests that the diploid must partially compensate for the loss of one CYH2 gene. Tetrad dissection shows that haploid spores lacking the CYH2 gene cannot germinate. The lethality of this deletion can be rescued by a CYH2 cDNA on a low copy vector. Haploids which lack the genomic copy of the CYH2 gene, but contain a plasmid copy of the CYH2 cDNA are able to grow normally. These CYH2 deleted yeast haploids should be useful to analyze mutationally altered CYH2 genes and genes homologous to CYH2 from other organisms without interference from a genomic copy.
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  • 78
    ISSN: 1432-0983
    Keywords: Yeast ; Gene regulation ; Saccharomyces cerevisiae ; PDCI promoter
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    Topics: Biology
    Notes: Summary A 870 by promoter fragment of the PDC1 gene that includes the carbon source dependent regulatory regions was investigated using 5′ and 3′ promoter deletions. The results indicate that glucose and ethanol regulation of PDC1 transcription are independently controlled by distinct cis-acting regions. The consensus sequence AAATCGATA may play a role in this regulation, while the sequence (ATCA)AACCT may be important in transcription initiation.
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  • 79
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    Current genetics 13 (1988), S. 21-23 
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Transformation ; Plasmid ; Colony ; Polyethylene glycol
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A rapid and simple yeast transformation procedure has been developed using colonies on agar plates. Saccharomyces cerevisiae SHY3 cells were picked up from colonies on YPD plates grown freshly or stored at 4 °C and incubated with M13RK9-T DNA at 30 °C for 1–2 h in a solution of Li+, Ca2+, Mg2+, triacetin and polyethylene glycol. About 3,500 transformants were obtained per µg of double stranded M13RK9-T DNA. Unlike the existing spheroplast techniques, single stranded M13RK9-T DNA transformed intact cells below one-hundredth frequency of the duplex form.
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  • 80
    ISSN: 1432-0983
    Keywords: Aspergillus terreus clonotheque ; Saccharomyces cerevisiae ; Homologous integration ; 2 μ circledirected chromosome destabilization
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    Notes: Summary A genome clonotheque consisting of 25- to 40-kb Sau3A1 fragments of Aspergillus terreus DNA was constructed in the episomal cosmid vector pES33 containing the yeastARG4 gene. From the 475 transformants of cir° yeast strain ESH-0, 23 stable Arg + transformants were independently selected. Genetic and Southern analysis of these stable transformants showed that 39% arose as a result of recombination between cloned A. terreus DNA sequences and yeast chromosome XII. The recombination events most likely occurred in the regions of homology within the rDNA clusters of A. terreus and Saccharomyces cerevisiae.
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  • 81
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    Current genetics 13 (1988), S. 283-289 
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Inducible antisense gene ; Acetolactate synthase ; Bradytrophic phenocopy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We report for the first time on the regulation of gene expression in yeast by antisense RNA. Chimaeric genes were constructed containing the 5′ upstream and partial coding sequence of SMR1 — a sulfometuron methyl resistant allele of the ILV2 locus. Such fragments were placed 5′ to 3′ and 3′ to 5′ under control of the GAL10 promoter and CYCl terminator in a high copy YEp plasmid. Following galactose induction only transformants containing antisense RNA genes showed biological activity against SMR1 gene expression. Antisense RNA inhibited synthesis of the SMR1 gene product acetolactate synthase and thus repressed cellular growth which resulted in a bradytrophic auxotroph revertable by addition of isoleucine and valine. Antisense RNA inhibition was enhanced in galactose medium containing sulfometuron methyl and in gcn4 cells deficient for positive regulation of the ILV2 locus. This system can be used to study factors that interfere with antisense RNA function and to assign biological function to randomly cloned DNA fragments.
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  • 82
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    Current genetics 14 (1988), S. 331-335 
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Meiosis ; Deletion mutations ; Sequence dissimilarities
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A diploid yeast strain with extensive sequence dissimilarity in homologous regions near the LYS2 locus was sporulated, and spontaneous lys2 and lys5 mutant spores, selected on α-amino adipate, were analyzed. As many as 50% of the mutant spores contained a deletion in LYS2. These deletions occurred at a frequency of 5.0 × 10−7. While deletions of various sizes and endpoints were obtained, all the deletions recovered in this study included the border between homologous and non-homologous sequences located 4 kb upstream of LYS2. Large lys2 deletions that extended into an adjacent CYH2 duplication occurred at a frequency of 2.0 × 10−7, more than 1,000 times the frequency of the CYH2-LYS2 deletions found in a related haploid strain. This high frequency of CYH2-LYS2 deletions was observed only after sporulation of the diploid strain, and was dependent upon extensive sequence dissimilarity near the LYS2 locus.
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  • 83
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Allosuppressor ; Translation ; Fidelity
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary Allosuppressor (sal) mutations enhance the efficiency of the yeast ochre suppressor SUQ5 and define five unlinked loci, SALT-SALS. A number of sal4 mutants were isolated and found to have pleiotropic, allele;specific phenotypes, including hypersensitivity in vivo to paromomycin and other antibiotics that stimulate translational errors in yeast. To examine further the nature of the SAL4 gene product, the wild type SAL4 gene was isolated by complementation of a conditional lethal allele sal4-2, and demonstrated to be a single copy gene encoding a single 1.6 kb transcript. Restriction mapping and DNA hybridisation analysis were used to demonstrate that the SAL4 gene is identical to the previously identified omnipotent suppressor gene SUP45 (SUPT). Our results implicate the SAL4 gene product as playing a major role in maintaining translational accuracy in yeast.
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  • 84
    ISSN: 1432-2048
    Keywords: Auxin (IAA), production by Rhizobium ; Gibberellin production by Rhizobium ; Mutant (Rhizobium) ; Nitrogen fixation ; Phaseolus (nodulation) ; Rhizobium (mutants) ; Root nodule
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Abstract Similar ranges of gibberellins (GAs) were detected by high-performance liquid chromatography (HPLC)-immunoassay procedures in ten cultures of wild-type and mutant strains of Rhizobium phaseoli. The major GAs excreted into the culture medium were GA1 and GA4. These identifications were confirmed by combined gas chromatographymass spectrometry. The HPLC-immunoassays also detected smaller amounts of GA9- as well as GA20-like compounds, the latter being present in some but not all cultures. In addition to GAs, all strains excreted indole-3-acetic acid (IAA) but there was no obvious relationship between the amounts of GA and IAA that accumulated. The Rhizobium strains studied included nod − and fix − mutants, making it unlikely that the IAA- and GA-biosynthesis genes are closely linked to the genes for nodulation and nitrogen fixation. The HPLC-immunoassay analyses showed also that nodules and non-nodulated roots of Phaseolus vulgaris L. contained similar spectra of GAs to R. phaseoli culture media. The GA pools in roots and nodules were of similar size, indicating that Rhizobium does not make a major contribution to the GA content of the infected tissue.
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  • 85
    ISSN: 1432-2048
    Keywords: Carbon dioxide fixation ; Citrulline ; Coralloid roots ; Cycads (nitrogen fixation) ; Nitrogen fixation ; Nitrogen transport ; Nostoc
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Freshly detached coralloid roots of several cycad species were found to bleed spontaneously from xylem, permitting identification of products of nitrogen transfer from symbiotic organ to host. Structural features relevant to the export of fixed N were described for Macrozamia riedlei (Fisch. ex Gaud.) Gardn. the principal species studied. Citrulline (Cit), glutamine (Gln) and glutamic acid (Glu), the latter usually in a lesser amount, were the principal translocated solutes in Macrozamia (5 spp.), Encephalartos (4 spp.) and Lepidozamia (1 sp.), while Gln and a smaller amount of Glu, but no Cit were present in xylem sap of Bowenia (1 sp.),and Cycas (2 spp.). Time-course studies of 15N enrichment of the different tissue zones and the xylem sap of 15N2-pulse-fed coralloid roots of M. riedlei showed earlier 15N incorporation into Gln than into Cit, and a subsequent net decline in the 15N of Gln of the coralloid-root tissues, whereas Cit labeling continued to increase in inner cortex and stele and in the xylem sap. Hydrolysis of the 15N-labeled Cit and Gln consistently demonstrated much more intense labeling of the respective carbamyl and amide groups than of the other N-atoms. Coralloid roots of M. riedlei pulse-fed 14CO2 in darkness showed 14C labeling of aspartic acid (Asp) and Cit in all tissue zones and of Cit of xylem bleeding sap. Lateral roots and uninfected apogeotropic roots of M. riedlei and M. moorei also incorporated 14CO2 into Cit. The 14C of Cit was restricted to the carbamyl-C. Comparable 15N2 and CO2-feeding studies on corallid roots of Cycas revoluta showed Gln to be the dominant product of N2 fixation, with Asp and alanine as other major 14C-labeled amino compounds, but a total absence of Cit in labeled or unlabeled form.
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  • 86
    ISSN: 1432-0983
    Keywords: Formaldehyde ; DNA-protein cross-links ; Repair ; Saccharomyces cerevisiae ; Hyperresistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The formation and removal of formaldehyde-mediated DNA protein cross-linking was measured by CsCI density gradient analysis in yeast strains of differing resistance to formaldehyde. Wild-type cells and transformants made hyperresistant to formaldehyde by a multi-copy vector containing the yeast SFA gene were specifically labeled in their DNA and incubated in the presence of formaldehyde. Treatment with formaldehyde lead to the formation of equal amounts of DNA protein cross-links; subsequent liquid holding of cells for 24 h resulted in the removal of nearly all DNA protein crosslinks regardless of the original formaldehyde resistance status of the strains.
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  • 87
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    Archives of microbiology 150 (1988), S. 326-332 
    ISSN: 1432-072X
    Keywords: Rhizobium leguminosarum ; Plasmids ; Melanin ; Nodulation ; Nitrogen fixation ; Plasmid curing
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Rhizobium leguminosarum strain VF39, isolated from nodules of field-grown faba beans in the Federal Republic of Germany, was shown to contain six plasmids ranging in molecular weight from 90 to 400 Md. Hybridisation to nif gene probes, plasmid curing, and mobilisation to other strains of Rhizobium and to Agrobacterium showed that the third largest plasmid, pRleVF39d (220 Md), carried genes for nodulation and nitrogen fixation. This plasmid was incompatible with pRL10JI, the Sym plasmid of R. leguminosarum strain JB300. Of the other plasmids, the two smallest (pRleVF39a and pRleVF39b, 90 and 160 Md respectively) were shown to be self-transmissible at a low frequency. Although melanin production is as yet unreported in strains of R. leguminosarum biovar viceae, strain VF39 produced a dark pigment, which, since it was not produced on minimal media and its production was greatly enhanced by the presence of tyrosine in the media, is probably melanin-like. Derivatives of VF39 cured of pRleVF39a no longer produced this pigment, but regained the ability to produce it when this plasmid was transferred into them. Strains of Agrobacterium tumefaciens, R. meliloti, and some strains of R. leguminosarum carrying pRleVF39a did not produce this pigment, indicating perhaps that some genes elsewhere on the VF39 genome are also involved in pigment production. Plasmid pRleVF39a appeared to be incompatible with the cryptic Rhizobium plasmids pRle336b and pRL8JI (both ca. 100 Md), but was compatible with the R. leguminosarum biovar phaseoli Sym plasmids pRP1JI, pRP2JI and pRph51a, all of which also code for melanin production. The absence of pRleVF39a in cured derivatives of VF39 had no effect on the symbiotic performance or competitive ability of this strain.
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  • 88
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    Archives of microbiology 150 (1988), S. 224-229 
    ISSN: 1432-072X
    Keywords: Vibrio ; V. diazotrophicus ; V. natriegens ; V. pelagius ; V. cincinnatiensis ; Nitrogenase ; Nitrogen fixation ; Oxygen sensitivity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Virtually all Vibrio spp. known and available in culture collections and several newly isolated Vibrio sp. were tested for their ability to fix molecular nitrogen, using the acetylene reduction technique, the fixation of the heavy isotope 15N, and by growth on media devoid of combined nitrogen. Among the 27 species tested, four, including V. diazotrophicus, proved to be nitrogenase-positive. The potential of nitrogen fixation was now also discovered in V. natriegens, V. pelagius and V. cincinnatiensis. Among the 9 newly isolated strains, 4 were nitrogenase-positive. These strains were classified as V. diazotrophicus on the basis of DNA homology studies. Nitrogenase was only induced during growth under anaerobic conditions. Dissolved oxygen as low as 1 μM inhibited nitrogenase completely. This inhibition at low oxygen concentration, however, was reversible. 50–100 μM dissolved oxygen inhibited nitrogenase irreversibly.
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  • 89
    ISSN: 1432-072X
    Keywords: Azorhizobium caulinodans ORS571 ; Hydrogenase ; Nitrogen fixation ; Chemostat cultures ; H2/N2 ratio ; ATP/2e value
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Hydrogenase-negative (Hup-) mutants of Azorhizobium caulinodans ORS571 were isolated by means of Tn5 mutagenesis. The colony test used for screening for Hup- strains was based on the absence of reduction of triphenyltetrazolium chloride with hydrogen. Suspensions from cultures of the mutant strains grown under derepressing conditions did not use hydrogen with methylene blue or oxygen as the hydrogen acceptor. The mutants were shown to carry single Tn5 insertions at different locations in the A. caulinodans genome. Molar growth yields (corrected for poly-β-hydroxybutyrate formation) in chemostat cultures of the mutants were similar to those of the wild type. Molar growth yields of the mutants were not increased by passing additional hydrogen through chemostat cultures, which is in agreement with the hydrogenase-negative phenotype of the mutants. H2/N2 ratios (mol H2 formed per mol N2 fixed) were calculated from the hydrogen content of the effluent gas and the N-content of the bacterial dry weight. Low H2/N2 ratios (between 1.2 and 1.9) were found in both energy-limited (oxygen or succinate) cultures and in cultures limited by the supply of an anabolic substrate (Mg2+). ATP/2e values (mol ATP used at the transport of 2e to nitrogen or H+) were calculated from the H2/N2 ratios and the molar growth yields of nitrogen-fixing and ammonia-assimilating cultures. ATP/2e values were between 7 and 11. It was concluded that the calculated ATP/2e values comprise not only 4 mol ATP used at the transport of 2e through nitrogenase but also energy equivalents needed for reversed electron flow from NADH to the low-potential hydrogen donor used by nitrogenase.
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  • 90
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    Archives of microbiology 151 (1988), S. 20-25 
    ISSN: 1432-072X
    Keywords: Yeast ; Saccharomyces cerevisiae ; Mating ; Zygote formation ; Chloroquine ; Lysosomotropic agent ; Plasma membrane ; Cell fusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Haploid cells of opposite mating type of Saccharomyces cerevisiae conjugate to form zygote. During the conjugation process, the degradation or reorganization of the cell wall and the fusion of the two plasma membranes take place. Since chloroquine inhibits cellular events associated with the reorganization of the plasma membrane, the effect of the drug on conjugation was studied. Chloroquine at a concentration, at which cell growth was not retarded, inhibited zygote formation, while it did not affect other mating functions, such as sexual agglutination, production of and response to mating pheromone. Cells in a mating culture containing chloroquine formed no “prezygote” suggesting that they were not prepared for entering into fusion process. The inhibitory effect of chloroquine was reversible as cells formed zygote when they were washed after treatment with chloroquine. Zygote formation was unaffected in cells possessing chlorquine within vacuoles after incubation with the drug in complete medium (YPD) at pH 7.5, followed by washing. This suggests that chloroquine inhibits zygote formaton by adsorbing to the plasma membrane of S. cerevisiae.
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  • 91
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    Archives of microbiology 150 (1988), S. 313-319 
    ISSN: 1432-072X
    Keywords: 3′-Phosphoadenylyl sulphate reductase ; Sulphite formation ; Cysteine biosynthesis ; Thioredoxin ; Saccharomyces cerevisiae ; HPLC enzyme analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The enzymatic mechanism of sulphite formation in Saccharomyces cerevisiae was investigated using a purified 3′-phosphoadenylsulphate (PAPS) reductase and thioredoxin. The functionally active protein (MR 80–85 k) is represented by a dimer which reduces 3′-phosphoadenylyl sulphate to adenosine-3′,5′-bisphosphate and free sulphite at a stoichiometry of 1:1. Reduced thioredoxin is required as cosubstrate. Examination of the reaction products showed that free anionic sulphite is formed with no evidence for “bound-sulphite(s)” as intermediate. V max of the enriched enzyme was 4–7 nmol sulphite · min-1 · mg-1 using the homologous thioredoxin from yeast. The velocity of reaction decreased to 0.4 nmol sulphite · min-1 · mg-1 when heterologous thioredoxin (from Escherichia coli) was used instead. The K m of homologous thioredoxin was 0.6 · 10-6 M, for the heterologous cosubstrate it increased to 1.4 · 10-6 M. The affinity for PAPS remained practically unaffected (K m PAPS: 19 · 10-6 M in the homologous, and 21 · 10-6 M in the heterologous system). From the kinetic data it is concluded that the enzyme followed an ordered mechanism with thioredoxin as first substrate followed by PAPS as the second. Parallel lines in the reciprocal and a common intersect in the Hanes-plots for thioredoxin were seen as indication of a ping-pong (with respect to thioredoxin) uni-bi (with respect to PAPS) mechanism.
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  • 92
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    Archives of microbiology 151 (1988), S. 44-48 
    ISSN: 1432-072X
    Keywords: Vanadium ; Molybdenum ; Methanogenesis ; Nitrogen fixation ; Archaebacterium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Nitrogen fixation of the Methanosarcina barkeri strains “Fusaro” (DSM 804) and “227” (DSM 1538) was found to be dependent on the presence of vanadium or molybdenum whereby molybdenum (added as Na2-molybdate) was preferred to vanadium (added as VCl3). Strain “227” showed less pronounced effects on diazotrophic growth with respect to vanadium and molybdenum. Rhenium (ReCl3) or tungsten (Na2-tungstate) could not replace vanadium or molybdenum. The optimum concentrations were found to be 2μM for vanadium and 5μM for molybdenum (strain “Fusaro”). This Mo optimum of methanogenesis was 10-fold higher with N2 than with NH4Cl as nitrogen source. A vanadium requirement with NH4Cl could not be detected. No interferences were observed if molybdenum and vanadium were added simultaneously under diazotrophic conditions. Growth yields were smallest for strain “227” grown diazotrophically ( $$Y_{CH_3 OH}$$ =0.6g dw/mol in the presence of vanadium and $$Y_{CH_3 OH}$$ =0.9g dw/mol in the presence of molybdenum), obviously higher for strain “Fusaro” grown diazotrophically ( $$Y_{CH_3 OH}$$ =1.15g dw/mol in the presence of V and $$Y_{CH_3 OH}$$ =1.4g dw/mol with Mo) and highest if M. barkeri was grown on NH4Cl as N-source ( $$Y_{CH_3 OH}$$ =3.4g dw/mol with Mo, strain “Fusaro”).
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  • 93
    ISSN: 1432-072X
    Keywords: Yeast ; Saccharomyces cerevisiae ; Mating reaction ; Zygote formation ; Mating pheromone ; Fatty acid ; Arachidonic acid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Effect of exogenous fatty acids on zygote formation in Saccharomyces cerevisiae was studied. Arachidonic and oleic acids considerably stimulated zygote formation, but other fatty acids tested, linoleic, linolenic, stearic and palmitic acids, did not. Pretreatment experiments with arachidonic acid showed that the stimulation of zygote formation by the fatty acid required the presence of mating pheromone.
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  • 94
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    Journal of industrial microbiology and biotechnology 2 (1988), S. 365-372 
    ISSN: 1476-5535
    Keywords: Osmotic pressure ; Intracellular ethanol ; Yeast ; Nutrient ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The intracellular accumulation of ethanol in yeast and its potential effects on growth and fermentation have been topics of controversy for the past several years. The determination of intracellular ethanol based on the exclusion of [14C]sorbitol to estimate aqueous cell volume was used to examine the question of intracellular ethanol accumulation. An intracellular accumulation of ethanol inSaccharomyces cerevisiae was observed during the early stages of fermentation. However, as fermentation continued, the intracellular and extracellular concentrations of ethanol became similar. Increasing the osmotic pressure of the medium with glucose or sorbitol was observed to cause an increase in the intracellular ethanol concentration. Associated with this was a decrease in yeast growth and fermentation rates. In addition, increasing the osmotic pressure of the medium was observed to cause an increase in glycerol production. Supplementation of the media with excess peptone, yeast extract, magnesium sulfate and potassium phosphate was found to relieve the detrimental effects of high osmotic pressure. Under these conditions, though, no effect on the intracellular and extracellular ethanol distribution was observed. These results indicate that nutrient limitation, and not necessarily intracellular ethanol accumulation, plays a key role during yeast fermentations in media of high osmolarity.
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  • 95
    ISSN: 1432-2048
    Keywords: Bacteroid ; Bradyrhizobium ; Glycine (N2 fixation) ; Nitrate reductase ; Nitrite reductase ; Nitrogen fixation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Bacteroids of Bradyrhizobium japonicum strain CB1809, unlike CC705, do not have a high level of constitutive nitrate reductase (NR; EC 1.7.99.4) in the soybean (Glycine max. Merr.) nodule. Ex planta both strains have a high activity of NR when cultured on 5 mM nitrate at 2% O2 (v/v). Nitrite reductase (NiR) was active in cultured cells of bradyrhizobia, but activity with succinate as electron donor was not detected in freshly-isolated bacteroids. A low activity was measured with reduced methyl viologen. When bacteroids of CC705 were incubated with nitrate there was a rapid production of nitrite which resulted in repression of NR. Subsequently when NiR was induced, nitrite was utilized and NR activity recovered. Nitrate reductase was induced in bacteroids of strain CB1809 when they were incubated in-vitro with nitrate or nitrite. Increase in NR activity was prevented by rifampicin (10 μg· ml-1) or chloramphenicol (50 μg·ml-1). Nitrite-reductase activity in bacteroids of strain CB1809 was induced in parallel with NR. When nitrate was supplied to soybeans nodulated with strain CC705, nitrite was detected in nodule extracts prepared in aqueous media and it accumulated during storage (1°C) and on further incubation at 25°C. Nitrite was not detected in nodule extracts prepared in ethanol. Thus nitrite accumulation in nodule tissue appears to occur only after maceration and although bacteroids of some strains of B. japonicum have a high level of a constitutive NR, they do not appear to reduce nitrate in the nodule because this anion does not gain access to the bacteroid zone. Soybeans nodulated with strains CC705 and CB1809 were equally sensitive to nitrate inhibition of N2 fixation.
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  • 96
    ISSN: 1432-1424
    Keywords: vacuole ; lipid bilayer ; K-channel ; single channel ; DIDS ; yeast ; Saccharomyces cerevisiae ; Ca2+ activation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary A voltage-dependent and Ca2+-activated cation channel found in the vacuolar membrane of the yeast,Saccharomyces cerevisiae, was incorporated into planar lipid bilayer and its gating characteristics were studied at the macroscopic and single-channel levels. The open-channel probability at steady state, which was estimated by the macroscopic current measurement, gave a maximum value at −10 mV and decreased in a graded fashion as the voltage became more positive or more negative. The steady-state voltage dependence was explained by assuming two independent gates, which had different rate constants and opposite voltage dependence. The fast-responding gate opened when the voltage of thecis side (the side to which the vesicles were added) was made more negative and the slow-responding gate behaved in the opposite direction. Relatively high concentrations of Ca2+, about 1mm, were required on thecis side for opening the slow gate in a voltage-dependent manner. DIDS increased the open-channel probability of the fast gate when added to thecis side, but was ineffective on the slow gate.
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  • 97
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    Molecular genetics and genomics 211 (1988), S. 88-94 
    ISSN: 1617-4623
    Keywords: CupI ; Gene amplification ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Carcinogen-induced amplification at the CupI locus, coding for a metallothionein protein, was studied in the yeast Saccharomyces cerevisiae. Exposure of cells from three different haploid strains, 4939, DBY746 and 320, to chemical carcinogens such as N-methyl-N′-nitro-N-nitrosoguanidine (MNNG), ethylmethanesulfonate (EMS) and 4-nitroquinoline-N-oxide (4NQO) enhanced the frequency of copper-resistant colonies up to several hundred fold. Copper-resistant clones obtained from strains DBY746 and 320, which contain more than one copy of the CupI locus, displayed a four-to eightfold amplification of the CupI sequences. In these clones the amplified CupI sequences were organized in a tandem array. Carcinogen treatment of strain 4939 in which only one copy of the CupI gene is present produced resistant colonies without CupI amplification. The possible use of the yeast system to study gene duplication and amplification is discussed.
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  • 98
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    Molecular genetics and genomics 211 (1988), S. 446-454 
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Invertase genes ; Promoter sequences ; Palindromes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary In the yeast Saccharomyces cerevisiae six unlinked structural genes for invertase, the SUC genes, are known. We sequenced about 800 bp of the 5′ non-coding region and the first 220 bp of the coding region of the genes SUC1, SUC3, SUC4 and SUC5 and compared them with the previously sequenced genes SUC2 and SUC7 (Sarokin and Carlson 1985a). All are highly homologous within the coding region but in the non-coding region SUC1 shows some differences and SUC2 is more highly diverged. Two different kinds of TATA boxes were identified: the more strongly expressed genes SUC1, 2 and 4 have the sequence TATAAA and the more weakly expressed genes SUC3, 5 and 7 have TACAAA. Though the SUC1 sequence is in general more homologous to the other SUC genes, the region between-140 and + 100 of SUC1 is nearly identical to SUC2. This could be due to a gene conversion between SUC1 and the silent suc2 o allele which occurs in the strains carrying SUC1. Within the upstream regions of all the SUC genes three regions with palindromic sequences analogous to stem and loop structures were identified. Comparable structure could be detected in similar positions in the upstream sequences of the divergently transcribed yeast gene pairs MAL6S-MAL6T and GAL1-GAL10. Implications for the importance of these structures in the regulation and initiation of transcription are discussed.
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  • 99
    ISSN: 1617-4623
    Keywords: l-azetidine-2-carboxylate resistance ; Escherichia coli ; γ-glutamyl kinase ; Proline ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We constructed plasmids carrying the Escherichia coli proB gene that encodes γ-glutamyl kinase, under the control of the yeast GAL1 promoter. This construction was carried out with both the wild-type proB + gene and a mutant allele, proB74, that specifies an enzyme resistant to feedback inhibition by proline. Yeast pro1 mutants harboring these plasmids are proline prototrophs. We conclude that the pro1 mutation results in a deficiency in the γ-glutamyl kinase activity in Saccharomyces cerevisiae. Expression of the proB74 allele in yeast resulted in enhanced resistance to the proline analogue l-azetidine-2-carboxylate and in a 2.4-fold elevation of the intracellular free proline levels. This result suggests that γ-glutamyl kinase is the rate limiting step in proline biosynthesis in yeast.
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  • 100
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    Molecular genetics and genomics 214 (1988), S. 249-256 
    ISSN: 1617-4623
    Keywords: Bent DNA ; DNA structure ; Saccharomyces cerevisiae ; 2 μm circle
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Until recently it was assumed that any short segment of DNA could be approximated as a straight rod. Many instances, however, have been reported in which the helical axis is curved. We have devised a simple method for selective identification of DNA segments containing a sequence-directed bend (curvature), by means of a two-dimensional polyacrylamide gel electrophoresis. In order to gain general insights into the structural features and the functional significance of sequence-directed bends, a bank of plasmids carrying bent DNA inserts from the Saccharomyces cerevisiae total genomic DNA was constructed. Primary characterizations of a set of bent DNA segments randomly cloned from S. cerevisiae are presented. One of the cloned DNA segments appears to be derived from a yeast plasmid, the 2 μm circle DNA.
    Type of Medium: Electronic Resource
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