Characteristics of an Arabidopsis glyoxylate reductase: general biochemical properties and substrate specificity for the recombinant protein, and developmental expression and implications for glyoxylate and succinic semialdehyde metabolism in planta
摘要：
Constitutive expression of an Arabidopsis thaliana (L.) Heynh cDNA (GenBank accession No. AY044183) in a succinic semialdehyde (SSA) dehydrogenase-deficient yeast (Saccharomyces cerevisiae Hansen) mutant enables growth on gamma-aminobutyrate and significantly enhances the accumulation of 7-hydroxybutyrate. In this report, the cDNA (designated hereinafter as AtGR1) was functionally expressed in Escherichia coli, and the reeombinant protein purified to homogeneity. Kinetic analysis of substrate specificity revealed that the enzyme catalyzed the conversion of glyoxylate to glycolate (K_(m, glyoxylate) = 4.5 mu mol-L~(-1)) as well as SSA to gamma-hydroxybutyrate (K_(m, SSA) = 0.87 mmol-L~(-1)) via an essentially irreversible, NADPH-based mechanism. Theenzyme had a 250-fold higher preference for glyoxylate than SSA based on the performance constants (k_(cat)/K_m), and with the exception of 4-carboxybenzaldehyde, at least a 100-fold higher preference for SSA than all other substrates tested (formaldehyde, acetaldehyde, butyraldehyde, 2-carboxybenzaldehyde, glyoxal, methyl-glyoxal, phenylglyoxal, phenylglyoxylate). In vitro assays of SSA reductase activity in cell-free extracts from Arabidopisis revealed its presence throughout the plant, although its specific activity was considerably higher in leaves at all developmental stages and in reproductive parts than in roots. It is proposed that the enzyme functions in redox homeostasis and the detoxification of both glyoxylate and SSA, in planta, resultingin the production of glycolate and 7-hydroxybutyrate, respectively.
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关键词：
aldo-keto reductase aldehyde reductase γ-aminobutyrate γ-hydroxybutyrate glyoxylate reductase succinic semialdehyde reductase
DOI：
10.1139/B07-081
被引量：
年份：
2007
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