caa a22 4500 605506124 CHVBK 20210128100625.0 cr unu---uuuuu 210128e20150701xx s 000 0 eng 10.1007/s00253-015-6597-y doi (NATIONALLICENCE)springer-10.1007/s00253-015-6597-y The pro-enzyme C-terminal processing domain of Pholiota nameko tyrosinase is responsible for folding of the N-terminal catalytic domain [Elektronische Daten] [Lai Moe, Saya Maekawa, Yasuko Kawamura-Konishi] Pholiota nameko (Pholiota microspore) tyrosinase is expressed as a latent 67-kDa pro-tyrosinase, comprising a 42-kDa N-terminal catalytic domain with a binuclear copper centre and a 25-kDa C-terminal domain and is activated by proteolytic digestion of the C-terminal domain. To investigate the role of the C-terminal processing domain of pro-tyrosinase, we constructed a recombinant tyrosinase lacking the C-terminal domain and four recombinant pro-tyrosinase mutants (F515G, H539N, L540G and Y543G) carrying substituted amino acid residues on the C-terminal domain. The recombinant tyrosinase lacking the C-terminal domain had no catalytic activity; whereas the mutant L540G was copper depleted, the other mutants had copper contents similar to that of the wild-type pro-tyrosinase. Proteolytic digestion activated the mutants H539N and Y543G following release of the C-terminal domain, and the resulting tyrosinases had higher K m values for t-butyl catechol than the wild-type pro-tyrosinase. The mutants F515G and L540G were degraded by proteolytic digestion and yielded smaller proteins with no activity. These data suggest that the C-terminal processing domain of P. nameko pro-tyrosinase is essential for correct folding of the N-terminal catalytic domain and acts as an intramolecular chaperone during assembly of the active-site conformation. Springer-Verlag Berlin Heidelberg, 2015 Pholiota nameko nationallicence Tyrosinase nationallicence Pro-enzyme nationallicence Processing nationallicence Chaperone nationallicence Moe Lai Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, 921-8836, Nonoichi, Ishikawa, Japan aut Maekawa Saya Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, 921-8836, Nonoichi, Ishikawa, Japan aut Kawamura-Konishi Yasuko Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, 921-8836, Nonoichi, Ishikawa, Japan aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/13(2015-07-01), 5499-5510 0175-7598 99:13<5499 2015 99 253 https://doi.org/10.1007/s00253-015-6597-y text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6597-y text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Moe Lai Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, 921-8836, Nonoichi, Ishikawa, Japan aut NATIONALLICENCE 700 1- Maekawa Saya Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, 921-8836, Nonoichi, Ishikawa, Japan aut NATIONALLICENCE 700 1- Kawamura-Konishi Yasuko Department of Food Science, Faculty of Bioresources and Environmental Sciences, Ishikawa Prefectural University, 1-308 Suematsu, 921-8836, Nonoichi, Ishikawa, Japan aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/13(2015-07-01), 5499-5510 0175-7598 99:13<5499 2015 99 253