Primary structure of tyrosinase from Neurospora crassa. II. Complete amino acid sequence and chemical structure of a tripeptide containing an unusual thioether.

To align the four cyanogen bromide peptides of Neurospora tyrosinase whose amino acid sequences were reported in the preceding paper, suitable methionine-containing overlap peptides were isolated. The required peptides were obtained by tryptic, peptic, and thermolytic digestion of the unmodified protein and of the maleylated derivative. From the partial sequence information of these peptides and a cyanogen bromide overlap peptide, the four cyanogen bromide fragments were aligned in the order CB3-CB1-CB4-CB2. These data establish Neurospora tyrosinase as a single-chain protein of 407 amino acids with a molecular weight of 46,000. The single cysteinyl residue 94 was found to be covalently linked via a thioether bridge to histidyl residue 96. The chemical nature of this unusual structure was elucidated by physicochemical analysis of peptides obtained from in vivo 35S, [2,5-3H]histidine, and [5-3H]histidine-labeled Neurospora tyrosinase.