Direct photoaffinity labeling of cysteine 211 or a nearby amino acid residue of beta-tubulin by guanosine 5 '-diphosphate bound in the exchangeable site

Tubulin with [8-C-14]GDP bound in the exchangeable site was exposed to ultr aviolet light, and radiolabel was cross-linked to two peptide regions of th e beta-subunit. Following enrichment for peptides cross-linked to guanosine by boronate chromatography, we confirmed that the cysteine 12 residue was the major site of crosslinking. However, significant radiolabel was also in corporated into a peptide containing amino acid residues 206 through 224. A lthough every amino acid in this peptide except cysteine 211 was identified by sequential Edman degradation, implying that this was the amino acid res idue cross-linked to guanosine, radiolabel at C-8 was usually lost during p eptide processing (probably during chromatography at pH 10). Consequently, the radiolabeled amino acid could not be unambiguously identified.