title: C-terminal processing of tyrosinase is responsible for activation of Pholiota microspora proenzyme.
abstract: Tyrosinase is expressed as a 67-kDa protein in Pholiota microspora (synonym Pholiota nameko), whereas the same enzyme purified from fruiting bodies of P. microspora is a 42-kDa protein that is cleaved with a C-terminal 25-kDa polypeptide from the 67-kDa protein. To confirm the role of C-terminal processing in enzyme activity, we expressed a recombinant 67-kDa tyrosinase in Escherichia coli cells. To obtain a soluble protein, the recombinant tyrosinase was expressed as a thioredoxin fusion protein with an enterokinase-cleavable site. Enterokinase digestion of the fusion protein produced a recombinant 67-kDa tyrosinase that did not have any catalytic activity. However, chymotrypsin digestion of the fusion protein produced a recombinant 44-kDa tyrosinase that was catalytically active and had a 25-kDa cleaved C-terminal. Kinetic parameters of the 44-kDa tyrosinase were similar to those of the 42-kDa tyrosinase purified from the fruiting bodies. These results suggest that tyrosinase is expressed in P. microspora as a latent 67-kDa proenzyme and is converted to the mature active 42-kDa enzyme by proteolytic processing of the C-terminal.
authors.affiliation: Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, Nonoichi, Ishikawa, 921-8836, Japan. konishi@ishikawa-pu.ac.jp
authors.forename: Yasuko
authors.initials: Y
authors.lastname: Kawamura-Konishi
authors.forename: Saya
authors.initials: S
authors.lastname: Maekawa
authors.forename: Mariko
authors.initials: M
authors.lastname: Tsuji
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Goto
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055368:Pholiota / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D011499:Protein Processing, Post-Translational
chemical_list: D004792:Enzyme Precursors; D005656:Fungal Proteins; D014442:Monophenol Monooxygenase