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  • 1
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 233 (2004), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: We have isolated putative esterase genes from various bacterial chromosomes. Thirty open reading frames predicted to encode esterases were randomly selected from 13 sequenced bacterial chromosomes and were cloned into an expression vector. The esterase activity of the resulting clones was tested on a tributyrin plate at different pH values and temperatures. Nine out of thirty tested clones exhibited significant tributyrin hydrolyzing activity. The enzyme S5 from the gene b0494 of Escherichia coli, the enzyme S12 from the gene STM0506 of Salmonella typhimurium, and the enzyme S28 from the gene AF1716 of Archaeoglobus fulgidus exhibited high activity at an alkaline pH range. The esterase S11 encoded by the gene PA3859 of Pseudomonas aeruginosa PAO1 and the esterase S21 from the gene SMc01033 of Sinorhizobium meliloti 1021, both showed a sharp increase in enzyme activity above pH 8.0. Furthermore, the enzymes S5, S12, S21, and S28 retained the esterase activity when they were incubated at 50 °C, suggesting that these enzymes are thermostable. Subsequent pH vs. activity and temperature vs. activity experiments with selected enzymes in a solution assay system confirmed the validity of the above data. The genome-wide exploration strategy of proteins provided valuable information on the esterases by revealing subtle biochemical differences between the esterases of different sources.
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 473-479 
    ISSN: 0006-3592
    Keywords: INU signal peptide ; MFα1 leader peptide ; secretion ; human lipocortin-1 ; human interleukin-2 ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The INU genes of Kluyveromyces marxianus encode inulinases which are readily secreted from Saccharomyces cerevisiae into the culture medium. To evaluate the utility of the INU signal peptides for the secretion of heterologous proteins from S. cerevisiae, a variety of expression and secretion vectors were constructed with GAL10 promoter and GAL7 terminator. The coding sequence for human lipocortin-1 (LC1) was inserted in-frame with the INU signal sequences, and then the secretion efficiency and localization of LC1 were investigated in more detail and compared with those when being expressed by the vector with the MFα1 leader peptide. The vector systems with INU signal peptides secreted ca. 95% of the total LC1 expressed into the extracellular medium, while the MFα1 leader peptide-containing vector resulted in very low secretion efficiency below 10%. In addition, recombinant human interleukin-2 (IL-2) was expressed and secreted with the vector systems with INU signal peptide, and a majority fraction of the human IL-2 expressed was found to be secreted into the extracellular medium as observed in LC1 expression. © 1995 John Wiley & Sons, Inc.
    Additional Material: 4 Ill.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 57 (1998), S. 245-249 
    ISSN: 0006-3592
    Keywords: human parathyroid hormone ; proteolysis ; L-arginine ; secretory production ; KEX2 endoproteinase ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A gene coding for human parathyroid hormone (hPTH) was synthesized and cloned into a yeast expression and secretion vector containing the mating factor α pre-pro leader sequence and the galactose-inducible promoter, GAL10. The intact hPTH(1-84) was found to be secreted into the culture medium. As observed in the previous reports on the secretory production of hPTH in yeast, however, the proteolytic cleavage occurred as the culture proceeded, resulting in a significant loss of the intact hPTH. Attempts were therefore made to reduce the extent of proteolysis by simply controlling the culture conditions. The proteolytic cleavage was significantly reduced by the addition of an excess amount of l-arginine (≥0.2M) to the culture medium, which resulted in a marked improvement in the yield of intact hPTH. To examine whether l-arginine affects the activities of intracellular proteases such as KEX2 endoproteinase or extracellular proteases, the proteolysis experiments were performed by incubating the commercial intact hPTH in a yeast host culture supernatant. The results demonstrated that l-arginine at high concentrations reduced the rate of hPTH proteolysis by inhibiting extracellular proteases. © 1998 John Wiley & Sons, Inc. Biotechnol. Bioeng. 57: 245-249, 1998.
    Additional Material: 4 Ill.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 29 (1987), S. 552-557 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A dual hollow fiber bioreactor, consisting of an outer silicone membrane for oxygen supply and an inner polyamide membrane for substrate permeation, was used as an immobilized enzyme reactor to carry out enzymatic glucose oxidation. Attaching a silicone tube oxygenator to provide an additional oxygen supply improved the conversion in glucose oxidation when the oxygen supply was rate-limiting. The reactor was operated in both diffusion and ultrafiltration modes. In the latter case, the conversion was much higher, but the stability of the immobilized enzyme was better maintained in the diffusion mode. As the inlet glucose concentration increased from 10mM to 500mM, the conversion decreased from 70 to 20%.
    Additional Material: 11 Ill.
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  • 5
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 32 (1988), S. 205-212 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Aspergillus niger B60 was immobilized in a dual hollow-fiber bioreactor (DHFBR) to produce citric acid continuously. The fungi proliferated well in the interstitial region formed by a parallel arrangement of three microporous polypropylene hollow fibers contained within a silicone tube. Long-term operation with nitrogen-enriched medium was not possible due to expansion of the silicone tubes by continual cell growth. The fungal growth could be controlled by supplying a nitrogen-deficient medium at the production stage. With pure oxygen aeration and nitrogen-deficient medium, volumetric productivity reached 1.62 g/L h at a residence time of 4.02 h, which corresponded to a 27-fold increase over that of shake-flask fermentation. When the residence time was increased to 20.1 h, citric acid at a concentration of 26 g/L was continuously produced, with a yield of 80-90% and a volumetric productivity of 1.3 g/L h. This represents a significant improvement in final concentration, yield, and the volumetric productivity over the equivalent values of the corresponding batch fermentation, which were 18 g/L, 40%, and 0.06 g/L h, respectively.
    Additional Material: 8 Ill.
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  • 6
    ISSN: 0749-503X
    Keywords: α1-antitrypsin ; Saccharomyces cerevisiae ; Hansenula polymorpha ; Pichia pastoris ; glycosylation ; secretion ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Human α1-antitrypsin (α1-AT) is a major serine protease inhibitor in plasma, secreted as a glycoprotein with a complex type of carbohydrate at three asparagine residues. To study glycosylation of heterologous proteins in yeast, we investigated the glycosylation pattern of the human α1-AT secreted in the baker's yeast Saccharomyces cerevisiae and in the methylotrophic yeasts, Hansenula polymorpha and Pichia pastoris. The partial digestion of the recombinant α1-AT with endoglycosidase H and the expression in the mnn9 deletion mutant of S. cerevisiae showed that the recombinant α1-AT secreted in S. cerevisiae was heterogeneous, consisting of molecules containing core carbohydrates on either two or all three asparagine residues. Besides the core carbohydrates, variable numbers of mannose outer chains were also added to some of the secreted α1-AT. The human α1-AT secreted in both methylotrophic yeasts was also heterogeneous and hypermannosylated as observed in S. cerevisiae, although the overall length of mannose outer chains of α1-AT in the methylotrophic yeasts appeared to be relatively shorter than those of α1-AT in S. cerevisiae. The α1-AT secreted from both methylotrophic yeasts retained its biological activity as an elastase inhibitor comparable to that of α1-AT from S. cerevisiae, suggesting that the different glycosylation profile does not affect the in vitro activity of the protein. © 1998 John Wiley & Sons, Ltd.
    Additional Material: 7 Ill.
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  • 7
    Publication Date: 2006-12-29
    Print ISSN: 1932-7447
    Electronic ISSN: 1932-7455
    Topics: Chemistry and Pharmacology
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  • 8
    Publication Date: 2015-06-05
    Print ISSN: 0175-7571
    Electronic ISSN: 1432-1017
    Topics: Biology , Physics
    Published by Springer
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  • 9
    Publication Date: 2006-05-25
    Print ISSN: 0175-7571
    Electronic ISSN: 1432-1017
    Topics: Biology , Physics
    Published by Springer
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  • 10
    Publication Date: 2009-03-17
    Print ISSN: 0743-7463
    Electronic ISSN: 1520-5827
    Topics: Chemistry and Pharmacology
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