Expression of α-L-arabinofuranosidase in Pichia pastoris and its effect on flterability of barley malt

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  • 1(The key Laboratory of Industrial Biotechnology,Jiangnan University,Wuxi 214122,China) 2(National Engineering Laboratory for Cereal Fermentation Technology,Jiangnan University,Wuxi 214122,China) 3(School of Biotechnology, Jiangnan University,Wuxi 214122,China) 4(Jiangsu Agribusiness Malt Co.Ltd,Sheyang 224300,China) 5( China Resources Snow Breweries Co.Ltd,Qinhuangdao 066001,China)

Online published: 2018-03-14

Abstract

The arabidofuranosidase gene was amplified from theAspergillus nigergenome by PCR. The gene length was 1790 bp containing a intron sequence with 50 bp. The corresponding protein sequence contains 9 N-glycosylation sites and 5 O-glycosylationsites, the N-terminal contains 18 amino acids of the signal peptide. Arabidofuranosidase gene was ligated with the expression vector pPICZαA and induced inPichia pastorX-33. The recombinant enzyme was purified by Ni column affinity chromatography. The molecular size of the recombinant enzyme is 70 kDa, and the optimum reaction temperature and optimum pH of the recombinant enzyme were 50 ℃ and 5.5 respectively. Cu2+, Zn2+and Fe2+had inhibitory effect on the activity of Arabidofuranosidase, Fe3+promoted the enzyme activity of arabidofuranosidase. TheKmandVmaxvalues of the enzymes were 0.78 mM and 2.57 μmol/min/mg respectively, using 4-Nitrophenyl α-L-arabinofuranoside as the substrate. Recombinant α-L-arabinofuranosidase was added at the initial stage of mashing with an amount of 31.2 mU/g, wort filtration rate was increased by 12.8%. α-L-arabinofuranosidase and xylanase have synergistic effect with barley malt arabinoxylans as substrate.

Cite this article

XIE Xi-zhu et al . Expression of α-L-arabinofuranosidase in Pichia pastoris and its effect on flterability of barley malt[J]. Food and Fermentation Industries, 2018 , 44(2) : 48 . DOI: 10.13995/j.cnki.11-1802/ts.015687

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