Characterization Of A ?-xylosidase And Its Synergistic Action On Biomass With Xylanase

With the fossil energy to be depleted,renewable resources are getting more attention than ever.Biomass resources are from various sources and multifunctional.As biotechnology advances fast,values of biomass resources have always been increasing.Lignocellulose is the most abundant biomass resource on earth.It can not only be converted into energy products,but also plenty of chemicals.Biotechnology plays an important role in the conversion.Hemicellulose is one of the three fractions of lignocellulosic biomass,accounting for a percentage of 20%-30%.While xylan,a kind of hemicellulose,is the second abundant biopolymer in the plant kingdom,after cellulose.The end product of hydrolysis of xylan is xylose,which can be utilized in different ways such as ethanol production by microbial fermentation.Both xylanase and ?-xylosidase are involved in the biodegradation of xylan.Xylanase cleaves the xylan into small fragments of polymers,while ?-xylosidase breaks down the fragments,freeing xylose.Thus,?-xylosidase plays an important role in the bioconversion of xylan to xylose.In this study,we implemented a research on the characterization of a ?-xylosidase and performed the synergy with a xylanase,XynA to degrade xylan.The ?-xylosidase gene?ThxylC?was cloned from Thermoanerobacterium aotearoense SCUT27,a species isolated from a hot spring in China.According to the amino acid sequence alignment,we identified the enzyme as a member of the GH120 family.ThXylC was tagged with a 6-histidine tag through DNA operation and expressed using E.coli BL21?DE3?and further purified by Ni-NTA affinity chromatography.ThXylC had a specific activity of 67.00 U/mg.The optimum temperature and pH were 65 oC and 6.5,respectively.ThXylC showed good tolerance against high temperature,withholding above 50 % activity after incubation at 65 oC for 48 h.Ki of ThXylC to xylose was 128 mM,which made it possible for further application in industry.Cu²⁺ and Ni²⁺ had strong inhibition on the hydrolysis activity and existence of 1 mM Cu²⁺ or Ni²⁺ inhibit the activity to 24.3 ± 4.4%? 70.1 ± 2.8%,respectively.Thin layer chromatography showed ThXylC could totally degrade oligoxylosaccharides?X2-X5?into xylose and positive synergy action of ThXylC and XynA?a xylanase?on hydrolysis of xylan.Test of the synergy effect to hydrolyze sugarcane bagasse showed that the produced reducing sugars increased from 0.233 to 0.366 mmol per gram sugarcane bagasse after the addition of ThXylC compared to that with XynA only.Synthetic research indicated that ThXylC has great potential for industrial application.