Biosynthetic activity of recombinant Escherichia coli-expressed Pichia etchellsii beta-glucosidase II.
The hydrolysis and biosynthetic reactions of partially purified Pichia etchellsii beta-glucosidase II from recombinant Escherichia coli pBG22:JM109 are described. With 167 mmol/L of initial glucose, the products of synthetic reactions, glucobiose and glucotriose, accumulated to 18 and 6 mmol/L, respectively. In transglycosylation reactions with 79 mmol/L of initial cellobiose, glucotriose and glucopentaose were obtained at 4.5 and 2 mmol/L, respectively. The effects of incubation time and substrate concentration were studied on the yield of synthesized oligosaccharides. In a reaction time of 24 h with 468 mmol/L of initial cellobiose, glucotriose and glucopentaose levels of 21.6 and 6.6 mmol/L, respectively, were obtained. The addition of dimethyl sulfoxide (DMSO) further increased the yields of the products by 10%. Detailed kinetic analysis indicated a significant (about twofold) increase in Vmax/KM of synthetic reactions in the presence of DMSO. A study of other disaccharides in transglycosylation reactions indicated biosynthetic activity in the order of sophorose > gentiobiose > cellobiose.
Bhatia
Yukti
Y
Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology, Delhi, Hauz Khas, New Delhi.
Mishra
Saroi
S
Bisaria
Virendra S
VS
D002475:Cellobiose / Q000378:metabolism; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004187:Disaccharides / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D005947:Glucose / Q000378:metabolism; D006031:Glycosylation; D006868:Hydrolysis; D007700:Kinetics; D010843:Pichia / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D001617:beta-Glucosidase / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism
D004187:Disaccharides; D011994:Recombinant Proteins; D002475:Cellobiose; D001617:beta-Glucosidase; D005947:Glucose; D004121:Dimethyl Sulfoxide