Glutamate‐Stimulated Phosphorylation of a Specific Protein in P2 Fractions of Rat Cerebral Cortex

Incubation of P2 fractions from rat cerebral cortex with 32Pi in the presence of L-glutamate caused an increased phosphorylation of a protein with apparent molecular weight of 43,000 (P43) as demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and autoradiography. This glutamate-stimulated phosphorylation of P43 was already detectable 10 s after the addition of glutamate and was dependent on the concentrations of glutamate in the incubation medium. Other excitatory amino acids such as D-glutamate, L-aspartate, D,L-cysteic acid, L-cysteinesulfinic acid, and D,L-alpha-aminoadipic acid did not stimulate the phosphorylation of P43. In contrast, alpha-ketoglutarate and succinate stimulated the phosphorylation of this protein. Glutamate-stimulated phosphorylation of P43 seemed not to be mediated by either cAMP or cGMP and was inhibited by the presence of Ca2+ in the incubation medium. Experiments performed with metabolic inhibitors indicated that glutamate-stimulated protein phosphorylation is localized in mitochondria. This conclusion is supported by the occurrence of glutamate-stimulated phosphorylation of P43 in mitochondrial fractions from several peripheral tissues. The present results are consistent with the hypothesis that P43 is a component of the pyruvate dehydrogenase complex of mitochondria.