Affinity labeling of adenine nucleotide-related enzymes with reactive adenine nucleotide analogs. II. Affinity labeling of phosphoglycerate kinase with a reactive AMP analog.

Affinity labeling of yeast and B. stearothermophilus phosphoglycerate kinases with a reactive AMP analog, N6-(p-bromoacetaminobenzyl)-AMP was examined. Complete loss of enzyme activity was observed when 1 mol of the reagent had reacted per mol of either enzyme. Results on the effect of pH and substrate addition on the inactivation, titration of SH groups before and after modification, and kinetic studies with AMP analogs suggest that the modification occurs at one amino group at or near the substrate binding site. General affinity labeling of kinases is discussed based on the results obtained.