A new mechanism of acyl group transfer in the reaction catalyzed by D-glyceraldehyde-3-phosphate dehydrogenase

D-Glyceraldehyde-3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12) catalyzes t he oxidative phosphorylation of its substrate in a two-step reaction. As a result of the first, oxidative step, the covalent intermediate wherein 3-ph osphoglyceroyl moiety is bound to Cys149 of the active center is subjected to nucleophilic attack by inorganic phosphate, but remains resistant to hyd rolytic decomposition. This ensures tight coupling of oxidation with phosph orylation in glycolysis. Ln this article, we present the experimental evide nce for the conversion of GAPDH into a form capable of performing the react ion in the absence of inorganic phosphate. The structural basis for this co nversion is the oxidation of a cysteine residue (probably Cys 153) into a s ulfenic acid derivative under mild conditions to affect the integrity of th e essential Cys 149. As a result, an intramolecular transfer of 3-phosphogl yceroyl group from the active center Cys 149 to Cys 153 becomes possible wi th subsequent hydrolysis of the sulfenyl carboxylate intermediate.