title: Cloning, expression, and characterization of two beta-glucosidases from isoflavone glycoside-hydrolyzing Bacillus subtilis natto.
abstract: On the basis of the genomic sequence of Bacillus subtilis 168, two beta-glucosidase genes (bglH and yckE) from B. subtilis natto, which has been reported to have high isoflavone glucoside-hydrolyzing activity, were cloned and overexpressed in E. coli M15. The temperature for the optimal p-nitrophenyl-beta-D-glucoside hydrolyzing activity of both enzymes was between 37 and 45 degrees C, but BglH had a higher thermal stability than YckE. Both showed high activity at pH 6.0, but YckE was stable over a wider pH range than BglH. Recombinant BglH was inhibited 73%, 63%, and 43% by 1.0 mM Cd(2+), Fe(2+), or Cu(2+), respectively, while other divalent metal ions resulted in 0-23% inhibition, whereas YckE was inhibited by less than 20% by any of the divalent metal ions we tested. Among the substrate we used, BglH showed the highest affinity for genistin and YckE showed the highest affinity for p-nitrophenyl-beta-D-fructopyranoside. Both BglH and YckE hydrolyzed genistin and daidzin into their isoflavone aglycones, genistein and daidzein, but BglH was more efficient than YckE in isoflavone glucoside hydrolysis (20-fold higher kcat). Our results suggest that recombinant BglH may be applicable in the process of isoflavones deglycosylation.
authors: Institute of Microbiology and Biochemistry, National Taiwan University, Taipei, Taiwan, ROC.
authors: Lun-Cheng
authors: LC
authors: Kuo
authors: Kung-Ta
authors: KT
authors: Lee
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology*; D044602:Cellulases / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D015870:Gene Expression; D006027:Glycosides / Q000378:metabolism; D006868:Hydrolysis; D007529:Isoflavones / Q000378:metabolism*; D007700:Kinetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D006027:Glycosides; D007529:Isoflavones; D011994:Recombinant Proteins; D044602:Cellulases