Production of Galactose Oxidase Inside the Fusarium Fujikuroi Species Complex and Recombinant Expression and Characterization of the Galactose Oxidase GaoA Protein from Fusarium Subglutinans

Overview

Journal Mol Biotechnol

Publisher Springer

Specialties Biotechnology
Molecular Biology

Date 2019 Jun 10

PMID 31177409

Citations 5

Authors

Carla Bertechini Faria

Fausto Fernandes de Castro

Damaris Batistao Martim

Camila Agnes Lumi Abe

Kelly Valerio Prates

Marco Aurelio Schuler de Oliveira

Ione Parra Barbosa-Tessmann

Affiliations

Soon will be listed here.

Abstract

Galactose oxidase catalyzes a two-electron oxidation, mainly from the C6 hydroxyl group of D-galactose, with the concomitant reduction of water to hydrogen peroxide. This enzyme is secreted by Fusarium species and has several biotechnological applications. In this study, a screening of galactose oxidase production among species of the Fusarium fujikuroi species complex demonstrated Fusarium subglutinans to be the main producer. The truncated F. subglutinans gaoA gene coding for the mature galactose oxidase was expressed from the prokaryotic vector pTrcHis2B in the E. coli Rosetta™ (DE3) strain. The purified recombinant enzyme presented temperature and pH optima of 30 °C and 7.0, respectively, K of 132.6 ± 18.18 mM, V of 3.2 ± 0.18 µmol of HO/min, k of 12,243 s, and a catalytic efficiency (k/K) of 9.2 × 10 M s. In the presence of 50% glycerol, the enzyme showed a T of 59.77 °C and was stable for several hours at pH 8.0 and 4 °C. Besides D-(+)-galactose, the purified enzyme also acted against D-(+)-raffinose, α-D-(+)-melibiose, and methyl-α-D-galactopyranoside, and was strongly inhibited by SDS. Although the F. subglutinans gaoA gene was successfully expressed in E. coli, its endogenous transcription was not confirmed by RT-PCR.

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