Abstract:

Glycoside hydrolase 61 family（GH61）belongs to the category of both oxidation and hydrolysis of cellulose degradation enzymes, it has a weak cellulose endonuclease, also can promote cellulase by oxidative damage to cellulose crystal structure on the degradation of lignocellulose, in the use of biomass resources it has potential application value. The GH61 family gene of Setosphaeria turcica was identified and bioinformatics to it was analyzed. Through transcriptome data analysis and fluorescence quantitative PCR verification, the GH61 family gene StGH61-11, induced by the carbon source corn stalk, was screened out. StGH61-11 was heterologous expressed, and its enzyme activity was determined by oxidation reaction with 2,6-dimethoxy-phenol, and the induction conditions were optimized. Its enzymatic properties were explored and the effect of StGH61-11 on promoting cellulase hydrolysis of cellulose was studied. The results showed that there were 21 GH61 family genes in S. turcica genome, and the filter paper enzyme activity of S. turcica significantly increased under the induction of corn stover. Transcriptome analysis of S. turcica showed that the expression levels of 11 genes in GH61 family increased when corn stover was used as carbon source. StGH61-11 was heterologous expressed in Escherichia coli. The optimal induction condition was 1 mmol/L IPTG at 25℃ for 9 h, and the specific activity reached（54.08±1.67）U/g. The optimal catalytic conditions were 50℃ and pH 5 for the recombinant protein StGH61-11 and the recombinant protein significantly improved the activity of cellulase hydrolysis of corn stover, with the highest degree of synergy up to 2.5 and the highest glycosylation rate up to 46.5%.

Key words: GH61 glycoside hydrolase family, lytic polysaccharide monooxygenases, cellulase, AA9, Setosphaeria turcica, synergistic degradation, heterologous expression