METABOLISM OF [3-C-13]PYRUVATE IN TCA CYCLE MUTANTS OF YEAST

The utilization of pyruvate and acetate by Saccharomyces cerevisiae was examined using C-13 and H-1 NMR methodology in intact wild-type yeast cells and mutant yeast cells lacking Krebs tricarboxylic acid (TCA) cycle enzymes. These mutant cells lacked either mitochondrial (NAD) isocitrate dehydrogenase (NAD-ICDH1), alpha-ketoglutarate dehydrogenase complex (alpha-KGDC), or mitochondrial malate dehydrogenase (MDH1). These mutant strains have the common phenotype of being unable to grow on acetate. [3-C-13]Pyruvate was utilized efficiently by wild-type yeast with the major intermediates being [C-13]glutamate, [C-13]acetate,and [C-13]alanine. Deletion of anyone of these Krebs TCA cycle enzymes changed the metabolic pattern such that the major synthetic product was [C-13]galactose instead of [C-13]glutamate, with some formation of [C-13]acetate and [C-13]alanine. The fact that glutamate formation did not occur readily in these mutants despite the metabolic capacity to synthesize glutamate from pyruvate is difficult to explain. We discuss the possibility that these data support the metabolon hypothesis of Krebs TCA cycle enzyme organization.