Počet záznamů: 1
Galactose Oxidase from Fusarium oxysporum - Expression in E. coli and P. pastoris and Biochemical Characterization
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SYSNO ASEP 0440666 Druh ASEP J - Článek v odborném periodiku Zařazení RIV J - Článek v odborném periodiku Poddruh J Článek ve WOS Název Galactose Oxidase from Fusarium oxysporum - Expression in E. coli and P. pastoris and Biochemical Characterization Tvůrce(i) Paukner, R. (AT)
Staudigl, P. (AT)
Choosri, W. (TH)
Sygmund, Ch. (AT)
Halada, Petr (MBU-M) RID, ORCID
Haltrich, D. (AT)
Leitner, CH. (AT)Celkový počet autorů 7 Zdroj.dok. PLoS ONE. - : Public Library of Science - ISSN 1932-6203
Roč. 9, č. 6 (2014)Poč.str. 8 s. Jazyk dok. eng - angličtina Země vyd. US - Spojené státy americké Klíč. slova galactose oxidase ; gene ; Fusarium ; gene expression Vědní obor RIV CE - Biochemie Institucionální podpora MBU-M - RVO:61388971 UT WOS 000338280800025 Anotace A gene coding for galactose 6-oxidase from Fusarium oxysporum G12 was cloned together with its native preprosequence and a C-terminal His-tag, and successfully expressed both in Escherichia coli and Pichia pastoris. The enzyme was subsequently purified and characterized. Among all tested substrates, the highest catalytic efficiency (k(cat)/K-m) was found with 1-methyl-beta-D-galactopyranoside (2.2 mM(-1) s(-1)). The Michaelis constant (K-m) for D-galactose was determined to be 47 mM. Optimal pH and temperature for the enzyme activity were 7.0 and 40 degrees C, respectively, and the enzyme was thermoinactivated at temperatures above 50 degrees C. GalOx contains a unique metalloradical complex consisting of a copper atom and a tyrosine residue covalently attached to the sulphur of a cysteine. The correct formation of this thioether bond during the heterologous expression in E. coli and P. pastoris could be unequivocally confirmed by MALDI mass spectrometry, which offers a convenient alternative to prove this Tyr-Cys crosslink, which is essential for the catalytic activity of GalOx. Pracoviště Mikrobiologický ústav Kontakt Eliška Spurná, eliska.spurna@biomed.cas.cz, Tel.: 241 062 231 Rok sběru 2015
Počet záznamů: 1