Fragmentation of proteins with o-iodosobenzoic acid: chemical mechanism and identification of o-iodoxybenzoic acid as a reactive contaminant that modifies tyrosyl residues

Abstract

O-Iodoxybenzoic acid, a disproportionation product of o-iodosobenzoic acid, was identified as a contaminant in most preparations of o-iodosobenzoic acid capable of both modifying and cleaving certain tyrosyl residues. A new synthetic approach for the production of o-iodosobenzoic acid containing low amounts of o-iodoxybenzoic acid combined with preincubation of the reagent with p-cresol to destroy the remaining o-iodoxybenzoic acid prior to the reaction with a polypeptide allows preparation of reagent solutions in which tyrosyl residues remain intact during tryptophanyl bond cleavage. In addition, the product produced by the action of o-iodosobenzoic acid upon tryptophanyl bonds was identified as N-acyldioxindolylalanine. It is inferred from that structure that the chemical reaction proceeds via a 2-step oxidation of the tryptophanyl residue followed by formation of an iminosporolactone which hydrolyzes, cleaving the peptide chain. Small peptides ending with dioxindolylalanine can be coupled to aminopropyl glass in high yield and are suitable for solid-phase Edman degradation.