Activation Mechanism of the Streptomyces Tyrosinase Assisted by the Caddie Protein

• Published in: Biochemistry (Easton) Vol. 56; no. 41; pp. 5593 - 5603
• Main Authors: Matoba, Yasuyuki, Kihara, Shogo, Muraki, Yoshimi, Bando, Naohiko, Yoshitsu, Hironari, Kuroda, Teruo, Sakaguchi, Miyuki, Kayama, Kure’e, Tai, Hulin, Hirota, Shun, Ogura, Takashi, Sugiyama, Masanori
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 17-10-2017
• Subjects: Amino Acid Substitution; Apoenzymes - chemistry; Apoenzymes - genetics; Apoenzymes - metabolism; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Benzoquinones - chemistry; Benzoquinones - metabolism; Binding Sites; Carrier Proteins - chemistry; Carrier Proteins - genetics; Carrier Proteins - metabolism; Catalytic Domain; Copper - chemistry; Copper - metabolism; Dihydroxyphenylalanine - analogs & derivatives; Dihydroxyphenylalanine - chemistry; Dihydroxyphenylalanine - metabolism; Enzyme Activation - drug effects; Models, Molecular; Monophenol Monooxygenase - chemistry; Monophenol Monooxygenase - genetics; Monophenol Monooxygenase - metabolism; Mutation; Oxidation-Reduction; Protein Aggregates - drug effects; Protein Multimerization - drug effects; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Reducing Agents - chemistry; Solubility; Streptomyces - enzymology; Tyrosine - chemistry; Tyrosine - metabolism
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/acs.biochem.7b00635

Tyrosinase (EC 1.14.18.1), which possesses two copper ions at the active center, catalyzes a rate-limiting reaction of melanogenesis, that is, the conversion of a phenol to the corresponding ortho-quinone. The enzyme from the genus Streptomyces is generated as a complex with a “caddie” protein that assists the transport of two copper ions into the active center. In this complex, the Tyr98 residue in the caddie protein was found to be accommodated in the pocket of the active center of tyrosinase, probably in a manner similar to that of l-tyrosine as a genuine substrate of tyrosinase. Under physiological conditions, the addition of the copper ion to the complex releases tyrosinase from the complex, in accordance with the aggregation of the caddie protein. The release of the copper-bound tyrosinase was found to be accelerated by adding reducing agents under aerobic conditions. Mass spectroscopic analysis indicated that the Tyr98 residue was converted to a reactive quinone, and resonance Raman spectroscopic analysis indicated that the conversion occurred through the formations of μ-η2:η2-peroxo-dicopper­(II) and Cu­(II)-semiquinone. Electron paramagnetic resonance analysis under anaerobic conditions and Fourier transform infrared spectroscopic analysis using CO as a structural probe under anaerobic conditions indicated that the copper transportation process to the active center is a reversible event in the tyrosinase/caddie complex. Aggregation of the caddie protein, which is triggered by the conversion of the Tyr98 residue to dopaquinone, may ensure the generation of fully activated tyrosinase.