Journal of Biological Chemistry
Volume 276, Issue 47, 23 November 2001, Pages 43775-43783
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ENZYME CATALYSIS AND REGULATION
NADP-Glutamate Dehydrogenase Isoenzymes of Saccharomyces cerevisiae: PURIFICATION, KINETIC PROPERTIES, AND PHYSIOLOGICAL ROLES*

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In the yeast Saccharomyces cerevisiae, two NADP+-dependent glutamate dehydrogenases (NADP-GDHs) encoded by GDH1 and GDH3 catalyze the synthesis of glutamate from ammonium and α-ketoglutarate. The GDH2-encoded NAD+-dependent glutamate dehydrogenase degrades glutamate producing ammonium and α-ketoglutarate. Until very recently, it was considered that only one biosynthetic NADP-GDH was present in S. cerevisiae. This fact hindered understanding the physiological role of each isoenzyme and the mechanisms involved in α-ketoglutarate channeling for glutamate biosynthesis. In this study, we purified and characterized the GDH1- and GDH3-encoded NADP-GDHs; they showed different allosteric properties and rates of α-ketoglutarate utilization. Analysis of the relative levels of these proteins revealed that the expression of GDH1 and GDH3 is differentially regulated and depends on the nature of the carbon source. Moreover, the physiological study of mutants lacking or overexpressing GDH1 or GDH3 suggested that these genes play nonredundant physiological roles. Our results indicate that the coordinated regulation of GDH1-, GDH3-, and GDH2-encoded enzymes results in glutamate biosynthesis and balanced utilization of α-ketoglutarate under fermentative and respiratory conditions. The possible relevance of the duplicated NADP-GDH pathway in the adaptation to facultative metabolism is discussed.

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Published, JBC Papers in Press, September 18, 2001, DOI 10.1074/jbc.M107986200

*

This work was supported in part by Dirección General de Asuntos del Personal Académico, Universidad Nacional Autónoma de México (UNAM), Grant IN212898 and by Consejo Nacional de Ciencia y Tecnologı́a Grant 31774.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Recipient of a fellowship and a grant (PAEP-102315) from the Dirección General de Estudios de Posgrado, UNAM.