Avendaño A, Deluna A, Olivera H, Valenzuela L, Gonzalez A
Departamento de Microbiología, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México City.
J Bacteriol. 1997 Sep;179(17):5594-7. doi: 10.1128/jb.179.17.5594-5597.1997.
It has been considered that the yeast Saccharomyces cerevisiae, like many other microorganisms, synthesizes glutamate through the action of NADP+-glutamate dehydrogenase (NADP+-GDH), encoded by GDH1, or through the combined action of glutamine synthetase and glutamate synthase (GOGAT), encoded by GLN1 and GLT1, respectively. A double mutant of S. cerevisiae lacking NADP+-GDH and GOGAT activities was constructed. This strain was able to grow on ammonium as the sole nitrogen source and thus to synthesize glutamate through an alternative pathway. A computer search for similarities between the GDH1 nucleotide sequence and the complete yeast genome was carried out. In addition to identifying its cognate sequence at chromosome XIV, the search found that GDH1 showed high identity with a previously recognized open reading frame (GDH3) of chromosome I. Triple mutants impaired in GDH1, GLT1, and GDH3 were obtained. These were strict glutamate auxotrophs. Our results indicate that GDH3 plays a significant physiological role, providing glutamate when GDH1 and GLT1 are impaired. This is the first example of a microorganism possessing three pathways for glutamate biosynthesis.
人们认为，酿酒酵母与许多其他微生物一样，通过由GDH1编码的NADP⁺-谷氨酸脱氢酶（NADP⁺-GDH）的作用，或通过分别由GLN1和GLT1编码的谷氨酰胺合成酶和谷氨酸合酶（GOGAT）的联合作用来合成谷氨酸。构建了缺乏NADP⁺-GDH和GOGAT活性的酿酒酵母双突变体。该菌株能够以铵作为唯一氮源生长，从而通过另一条途径合成谷氨酸。对GDH1核苷酸序列与完整酵母基因组之间的相似性进行了计算机搜索。除了在第十四条染色体上鉴定出其同源序列外，搜索还发现GDH1与第一条染色体上先前识别的开放阅读框（GDH3）具有高度同一性。获得了在GDH1、GLT1和GDH3中受损的三突变体。这些是严格的谷氨酸营养缺陷型。我们的结果表明，GDH3发挥着重要的生理作用，在GDH1和GLT1受损时提供谷氨酸。这是微生物拥有三种谷氨酸生物合成途径的第一个例子。
