The Reactivity of a Functional Tyrosyl Residue in Carboxypeptidase B: Nitration of the Cadmium Enzyme

Cadmium‐carboxypeptidase B was nitrated with tetranitromethane. The enzyme polymerized extensively during nitration. In the monomer nitrated Cd‐carboxypeptidase B, 70% of the activity of Cd‐carboxypeptidase B was retained. In order to identify the tyrosyl residues nitrated, the enzyme was digested with chymotrypsin and subtilisin and the nitrotyrosyl peptides were purified by affinity chromatography on antityrosyl‐antibody‐Sepharose conjugate followed by two‐dimensional thin‐layer chromatography. The major nitropeptides, representing 65% of the nitrotyrosyl label, were compatible with the segment of the sequence containing Tyr‐240 and Tyr‐259. Only 10% of the nitrotyrosyl label was found in the segment of Tyr‐248. This indicates that the state of Tyr‐248 in Cd‐carboxypeptidase B differs from that in zinc‐carboxypeptidase B where it shows chemical hyperreactivity due to its proximity to the metal ion.