ESSENTIAL TYROSINE RESIDUES IN TRANSKETOLASE OF BAKERS-YEAST .1. DETERMINATION OF THE TYPE AND NUMBER OF ESSENTIAL RESIDUES

Transketolase is completely and irreversibly inactivated in the presen ce of N-acetylimidazole, 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-C l), or tetranitromethane (TNM). The pH dependence of rare constants wh ich characterize TNM- or N-acetylimidazole-induced transketolase inact ivation indicates that the agents modify, tyrosine residues. Changes i n spectral properties caused by transketolase nitration are suggestive of nitrotyrosine accumulation. The activity of transketolase modified with N-acetylimidazole or NBD-Cl is substantially increased following selective displacement of the modifiers from tyrosine residues with h ydroxylamine and sulfhydryl reagents, respectively. Analysis of the co rrelation between the residual activity of the partially modified enzy me and the proportion of unmodified tyrosine residues (the Tsou method ) shows that transketolase contains at least 2 essential tyrosine resi dues per monomer.