Characterization of the two saccharopine dehydrogenase isozymes of lysine catabolism encoded by the single composite AtLKR/SDH locus of Arabidopsis.
Arabidopsis plants possess a composite AtLKR/SDH locus encoding two different polypeptides involved in lysine catabolism: a bifunctional lysine-ketoglutarate reductase/saccharopine dehydrogenase (LKR/SDH) enzyme and a monofunctional SDH enzyme. To unravel the physiological significance of these two enzymes, we analyzed their subcellular localization and detailed biochemical properties. Sucrose gradient analysis showed that the two enzymes are localized in the cytosol and therefore may operate at relatively neutral pH values in vivo. Yet while the physiological pH may provide an optimum environment for LKR activity, the pH optima for the activities of both the linked and non-linked SDH enzymes were above pH 9, suggesting that these two enzymes may operate under suboptimal conditions in vivo. The basic biochemical properties of the monofunctional SDH, including its pH optimum as well as the apparent Michaelis constant (K(m)) values for its substrates saccharopine and nicotinamide adenine dinucleotide at neutral and basic pH values, were similar to those of its SDH counterpart that is linked to LKR. Taken together, our results suggest that production of the monofunctional SDH provides Arabidopsis plants with enhanced levels of SDH activity (maximum initial velocity), rather than with an SDH isozyme with significantly altered kinetic parameters. Excess levels of this enzyme might enable efficient flux of lysine catabolism via the SDH reaction in the unfavorable physiological pH of the cytosol.
Zhu
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Department of Plant Sciences, The Weizmann Institute of Science, Rehovot 76100, Israel.
Tang
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G
Galili
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D017360:Arabidopsis / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D015153:Blotting, Western; D002458:Cell Fractionation; D003600:Cytosol / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes; D008239:Lysine / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D012443:Saccharopine Dehydrogenases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D007527:Isoenzymes; D010940:Plant Proteins; D012443:Saccharopine Dehydrogenases; C029496:saccharopine dehydrogenase (NADP+, lysine-forming); D008239:Lysine