Location of Histidine Residues that React with Bromoacetazolamide and N-Bromoacetylacetazolamide in Human and Bovine Carbonic Anhydrases

Abstract

Human carbonic anhydrase C and bovine carbonic anhydrase B were modified with the affinity label, bromoacetazolamide, and human carbonic anhydrase B with N-bromoacetylacetazolamide. Tryptic peptides from the modified enzymes, containing alkylated histidines, have been isolated by ion exchange and two-dimensional high voltage electrophoresis-paper chromatography. The partial amino acid sequence of the peptide from alkylated bovine enzyme B has been found to be Met-Val-Asn-Asn-Gly-His-Ser-Phe-Asn-Val-Glu-Tyr-Asx-Asx(Glx,Asx,Ser)Lys-. In all probability the histidine in this sequence that reacted is His-64. Comparison of the amino acid composition of the tryptic peptides from the two human isoenzymes with the proposed sequences of these enzymes has made it possible to identify histidine-64 in human enzyme C as that reacting with bromoacetazolamide and histidine-67 in human enzyme B as that reacting with N-bromoacetylacetazolamide.