Oxidation of an Adjacent Methionine Residue Inhibits Regulatory Seryl-Phosphorylation of Pyruvate Dehydrogenase:

A Met residue is located adjacent to phosphorylation site 1 in the sequences of mitochondrial pyruvate dehydrogenase E1α subunits. When synthetic peptides including site 1 were treated with H2O2, the Met residue was oxidized to methionine sulfoxide (MetSO), and the peptides were no longer phosphorylated by E1α-kinase. Isolated mitochondria were incubated under state III or IV conditions, lysed, the pyruvate dehydrogenase complex (PDC) immunoprecipitated, and tryptic peptides analyzed by MALDI-TOF mass spectrometry. In all instances both Met and MetSO site 1 tryptic-peptides were detected. Similar results were obtained when suspension-cultured cells were incubated with chemical agents known to stimulate production of reactive oxygen species within the mitochondria. Treatment with these agents had no effect upon the amount of total PDC, but decreased the proportion of P-PDC. We propose that the redox-state of the Met residue adjacent to phosphorylation site 1 of pyruvate dehydrogenase contributes to overall regulation of PDC activity in vivo.

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