Study On The Enzymatic Properties Of Ferulic Acid Esterase From Lactobacillus Plantarum And Its Application In Wheat Bran Fermentation

Ferulic acid Esterase(FAE)can hydrolyze the ester bond between ferulic acid and polysaccharide polymers to separate ferulic acid,assist other cell wall degrading enzymes to degrade plant cell walls,and separate a variety of phenolic acid which beneficial to human body.In this paper,ferulic esterase derived from multiple Lactobacillus plantarum was investigated.In view of the enzymatic characteristics of ferulic esterase of Lactobacillus plantarum and the efficiency of ferulic esterase in degrading wheat bran to produce ferulic acid,the following studies were conducted:(1)In this paper,primers were designed based on the template the ferulic acid esterase gene lp?0796 drived from the Lactobacillus plantarum WCFS1 in the NCBI database,and then ten Lactobacillus plantarums which save in laboratory were homologously cloned.The FAE gene sequences were used to construct recombinant plasmid,and four recombinant FAE plasmid vectors were successful builded:pET28A-FAE24,pET28A-FAE136/203,pET28A-FAE285,pET28A-FAE287.Which were transformed into E.coli for expression of the target protein.After purification by a dextran chromatography column,the purified FAE target protein was obtained according to the peak position.(2)The enzyme properties of the purified four recombinant FAE were studied,and them were determined that the optimal pH value was 7,and the optimal temperature was all 25?.After the enzymatic reaction kinetic analysis was performed under the optimal environment,it was found that the K_mvalue of pET28A-FAE287 was 1.181 mmol/L,the K_catvalue was 6.296 min,and the K_cat/K_mvalue was 5.331L/min.The enzyme activity of FAE287 was much higher than that of the other three recombinant FAE.Therefore,this enzyme was selected for subsequent wheat bran transformation experiment.(3)Under the optimal conditions[pH7,25?],pET28A-FAE287 could not hydrolyze wheat bran alone to release ferulic acid,but it could hydrolyze wheat bran with xylanase to produce 0.30 mg/g ferulic acid.Compared with the direct alkali hydrolysis of wheat bran,it accounted for 19.74%of the yield.At the same time,the hydrolysis of starch-removed wheat bran with xylanase could produce 0.40 mg/g ferulic acid,which accounts for12.23%of the alkali-hydrolysis of starch-removed wheat bran.This study provides a theoretical basis for the industrial transformation of wheat bran with ferulate esterase derived from Lactobacillus plantarum.