Study On The Structure And Function Of Beta-glucosidase From Enterococcus Faecalis

As one of the most abundant renewable energy sources on earth,plant biomass is mainly composed of lignocellulose.Lignocellulose is a complex heterogeneous complex composed of hemicellulose,lignin,and cellulose.In industry,cellulose can be degraded into fiber disaccharide as raw material,and widely used in biofuels,food,environmental protection,and other industries.How to degrade cellulose to produce glucose at low cost and high efficiency is the main bottleneck of the technology.At present,the main method of cellulose degradation is enzyme method.There are three kinds of enzymes:endoglucanase,fibrinolysins hydrolase,and?-glucosidase.The degradation of cellulose is mainly due to the break of?-1,4 glycoside bond,which is not catalyzed by many?-glycosidases in nature.Therefore,it is very important to screen and separate glycosidases which can catalyze the?-1,4-glycoside bond from nature.In the previous study,the team isolated a gram-positive strain that could degrade cellulose from the gut of pigs.This strain belongs to Enterococcus faecalis,which is a probiotic bacterium in the gut of human and most animals.6-phosphate-?-glucosidase was successfully amplified from the strain,belonging to GH1 family.In vitro,the recombinant expression system of E.coli was successfully expressed and purified by Ni column,ion column and molecular sieve to obtain high purity 6-phosphate-?-glucosidase.The results of enzyme-promoted experiments showed that the enzyme could catalyze the production of glucose from cellobiose,4-nitro-?-D-pyranogalactoside,p NPC and p-nitrophenyl-?-D-glucoside.Among them,the K_mvalues of 4-nitro-?-D-galactosidase and p-nitrophenyl-?-D-glucoside are the most suitable substrate.The optimal p H of 6-phosphate-?-glucosidase is 8.5-9.0 and the optimum temperature is between 40-60?,if Mg²⁺is added to the enzyme reaction process,the reaction rate will be increased,while Mn²⁺or Ni²⁺will inhibit the enzyme activity.To understand the catalytic mechanism of the enzyme,we used the structural biology method to study the crystallography of the protease and screen the crystal in index I test kit purchased by Hamptons.The results provide a basis for further analysis of its spatial structure and the structure of the enzyme and the complexes of different substrates.