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  • 1
    ISSN: 1432-2048
    Keywords: Brassica ; Embryo ; Carbon partitioning ; Fatty acid synthesis ; Plastid ; Starch synthesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The aim of this work was to investigate the partitioning of imported glucose 6-phosphate (Glc6P) to starch and fatty acids, and to CO2 via the oxidative pentose phosphate pathway (OPPP) in plastids isolated from developing embryos of oilseed rape (Brassica napus L.). The ability of the isolated plastids to utilize concurrently supplied substrates and the effects of these substrate combinations on the Glc6P partitioning were also assessed. The relative fluxes of carbon from Glc6P to starch, fatty acids, and to CO2 via the OPPP were close to 2∶1∶1 when Glc6P was supplied alone. Under these conditions NADPH generated via the OPPP was greater than that required by the concurrent rate of fatty acid synthesis. Fatty acid synthesis was unaffected by the presence or absence of exogenous NADH and/or NADPH and the requirement of fatty acid synthesis for reducing power is therefore met entirely by intraplastidial metabolism. When Glc6P was supplied in the presence of either pyruvate or pyruvate and acetate, the total flux from these metabolites to fatty acids was up to threefold greater than that from either Glc6P or pyruvate when they were supplied singly. In these experiments there was little competition between Glc6P and pyruvate in fatty acid synthesis and the flux to starch was unchanged. This implies that the starch and fatty acid biosynthesis pathways did not compete for the exogenously supplied ATP on which they were strongly dependent. When Glc6P and pyruvate were provided together, the NADPH generated by the OPPP pathway was less than that required by the concurrent rate of fatty acid synthesis. This suggests that the metabolism of exogenous Glc6P via the OPPP can contribute to the NADPH demand created during fatty acid synthesis but it also indicates that other intraplastidial sources of reducing power must be available under the in-vitro conditions used.
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  • 2
    ISSN: 1432-2048
    Keywords: Acetyl-CoA carboxylase ; Brassica ; Embryo (development) ; Fatty acid synthesis ; Pisum ; Ricinus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) activity has been determined in seed tissues of oilseed rape (Brassica napus L.), pea (Pisum sativum L.) and castor bean (Ricinus communis L.). A new method is described which leads to significantly higher measurable activities of the enzyme in tissue homogenates than previously reported. This method does not involve either Triton X-100 or centrifugation treatments which have been used previously in the study of the enzyme. In the case of oilseed rape the activity was also increased by removal of the testa from the seed. The activity of ACCase was determined throughout the development of oilseed rape embryos. Enzyme activity increased 3.5-fold as the embryo fresh weight increased from 0.3 to 2.0 mg and then reached a plateau at 1.1 nmol malonyl-CoA-min−1 · embryo−1. The main period of lipid accumulation commenced at an embryo fresh weight of 2.3 mg, which was after the plateau in ACCase activity had been reached. Activity of the enzyme declined after an embryo fresh weight of 3.5 mg, which was before lipid accumulation in the embryo had been completed. Comparison of the activity of ACCase and the apparent in-vivo rate of lipid synthesis on an embryo-fresh-weight basis (i.e. nmol malonyl-CoA formed or utilized·min−1·mg−1 fresh weight) revealed that ACCase activity declines relative to the rate of lipid synthesis throughout development. The negative correlation between these two rates is discussed in relation to the role of ACCase in the regulation of accumulation of storage lipid during embryo development.
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  • 3
    ISSN: 1432-2048
    Keywords: Key words: Amylase (isoforms) ; Brassica (starch metabolism) ; Embryo (developing) ; Plastid ; Starch phosphorylase (isoforms) ; Starch (synthesis/degradation)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract. The aim of this work was to characterise the metabolism of starch in developing embryos of oilseed rape (Brassica napus L. cv. Topaz). The accumulation of starch in embryos in siliques which were darkened or had been exposed to the light was similar, suggesting that the starch is synthesised from imported sucrose rather than via photosynthesis in the embryo. Starch content and the activities of plastidial enzymes required for synthesis of starch from glucose 6-phosphate (Glc6P) both peaked during the early-mid stage of cotyledon development (i.e. during the early part of oil accumulation) and then declined. The mature embryo contained almost no starch. The starch-degrading enzymes α-(EC 3.2.1.1) and β-amylase (EC 3.2.1.2) and phosphorylase (EC 2.4.1.1) were present throughout development. Most of the activity of these three enzymes was extraplastidial and therefore unlikely to be involved in starch degradation, but there were distinct plastidial and extraplastidial isoforms of all three enzymes. Activity gels indicated that distinct plastidial isoforms increase during the change from net synthesis to net degradation of starch. Plastids isolated from embryos at stages both before and after the maximum starch content could convert Glc6P to starch although the rate was lower at the later stage. The results are consistent with the idea that starch synthesis and degradation occur simultaneously during embryo development. The possible roles of transient starch accumulation during embryo development are discussed.
    Type of Medium: Electronic Resource
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