Inactivation of ribulosebisphosphate carboxylase by modification of arginyl residues with phenylglyoxal

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Abstract
Phenylglyoxal rapidly and completely inactivates spinach and Rhodospirillum rubrum ribulosebisphosphate carboxylases [EC 4.1.1.39]. Inactivation exhibits pseudo-1st-order kinetics and a reaction order of approximately 1 for both enzymes, suggesting that modification of a single residue per protomeric unit suffices for inactivation. Loss of enzymic activity is directly proportional to incorporation of [14C]phenylglyoxal until only 30% of the initial activity remains. For both enzymes, extrapolation of incorporation to 100% inactivation yields 4-5 mol of [14C]phenylglyoxal/mol protomer. Amino acid analyses confirm the expected 2:1 stoichiometry between phenylglyoxal incorporation and arginyl modification and suggest that other kinds of amino acid residues are not modified. (Thus, inactivation correlates with modification of 2-3 arginyl residues/protomer). The substrate ribulose bisphosphate and some competitive inhibitors reduce the rates of inactivation of the carboxylases and prevent modification of about 0.5-1.0 arginyl residue/protomer. Inactivation is therefore a consequence of modification of a small number of residues of the 35 and 29 total arginyl residues/protomer in spinach and R. rubrum carboxylases, respectively.