Multifunctions of MelB, a Fungal Tyrosinase from Aspergillus oryzae

• Published in: Chembiochem : a European journal of chemical biology Vol. 13; no. 2; pp. 193 - 201
• Main Authors: Fujieda, Nobutaka, Murata, Michiaki, Yabuta, Shintaro, Ikeda, Takuya, Shimokawa, Chizu, Nakamura, Yukihiro, Hata, Yoji, Itoh, Shinobu
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 23.01.2012; WILEY‐VCH Verlag
• Subjects: Amino Acid Sequence; Animals; Aspergillus oryzae - enzymology; copper; Dimerization; dioxygen activation; Electrophoresis, Polyacrylamide Gel; Enzyme Activation; enzyme function; hemocyanin; Models, Molecular; Molecular Sequence Data; Mollusca - chemistry; Mollusca - enzymology; Mollusca - genetics; Monophenol Monooxygenase - chemistry; Monophenol Monooxygenase - genetics; Monophenol Monooxygenase - metabolism; proenzymes; Sequence Alignment; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; tyrosinases
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.201100609

The pro form of melB tyrosinase from the melB gene of Aspergillus oryzae was over‐produced from E. coli and formed a homodimer that exhibited the spectral features of met‐tyrosinase. In the presence of NH2OH (reductant), the proenzyme bound dioxygen to give a stable (μ‐η2:η2‐peroxo)dicopper(II) species (oxy form), thus indicating that the pro form tyrosinase can function as an oxygen carrier or storage protein like hemocyanin. The pro form tyrosinase itself showed no catalytic activity toward external substrates, but proteolytic digestion with trypsin activated it to induce tyrosinase activity. Mass spectroscopy analyses, mutagenesis experiments, and colorimetry assays have demonstrated that the tryptic digestion induced cleavage of the C‐terminal domain (Glu458–Ala616), although the dimeric structure of the enzyme was retained. The structural changes induced by proteolytic digestion might open the entrance to the enzyme active site for substrate incorporation. Proteolytic functional change: the melB gene product (melB tyrosinase) from Aspergillus oryzae can function as dioxygen carrier, monooxygenase, and oxidase. The cleavage of the C‐terminal domain by proteolytic treatment changes the function of the tyrosinase from dioxygen carrier to catechol oxidase and phenol monooxygenase.