New substrates of 5-oxo-L-prolinase.

Abstract

Interaction of 5-oxo-L-prolinase (which catalyzes stoichiometric cleavage of 5-oxo-L-proline to L-glutamate coupled to that of cleavage of ATP to ADP and Pi), with a number of new 5-oxo-L-proline analogs was examined in studies in which (a) analogs were substituted for 5-oxo-L-proline and the formation of products was determined, (b) the effects of the analogs on the normal catalytic reaction were observed, and (c) the effects of the analogs on the ITPase activity exhibited by the enzyme were examined. Some of these reactions are partially coupled, i.e. the molar formation of ADP exceeds that of amino acid; others are uncoupled, i.e. cleavage of ATP, but not that of imino acid occurs. Analogs in which the 4-methylene moiety of 5-oxo-L-proline is replaced by O, S, and NH (L-2-oxooxazolidine-4-carboxylate, L-2-oxothiazolidine-4-carboxylate, and 2-imidazolidone-4-carboxylate) participate in reactions that are, partially coupled, coupled, and uncoupled respectively. Partial coupling occurs with cis- and trans-2-oxo-5-methyloxazolidine-4-carboxylate, and uncoupled reactions occur with L-2-iminothiazolidine-4-carboxylate, 2-thiazolinone-4-carboxylate, and 2-oxo-5,5-dimethylthiazolidine-4-carboxylate. These and earlier findings indicate that significant binding of the imino acid substrate requires a 5-carbonyl (or = NH), an unsubstituted N-1 and a C-2 of the L-configuration; substantial modification of 5-oxo-L-proline in the region of C-3 and C-4 is possible with retention of binding properties. Uncoupling and partial coupling may be associated with significant differences in the orientations of the analogs at the active site that facilitate (to various extents) the access of water to the nucleoside triphosphate.