Enzymatic synthesis of alpha-glucosyl-timosaponin BII catalyzed by the extremely thermophilic enzyme: Toruzyme 3.0L.
Timosaponin BII (BII), a steroidal saponin showing potential anti-dementia activity, was converted into its glucosylation derivatives by Toruzyme 3.0L. Nine products with different degrees of glucosylation were purified and their structures were elucidated on the basis of (13)C NMR, HR-ESI-MS, and FAB-MS spectra data. The active enzyme in Toruzyme 3.0L was purified to electrophoretic homogeneity by tracking BII-glycosylase activity and was identified as Cyclodextrin-glycosyltransferase (CGTase, EC 2.4.1.19) by ESI-Q-TOF MS/MS. In this work, we found that the active enzyme catalyzed the synthesis of alpha-(1-->4)-linked glucosyl-BII when dextrin instead of an expensive activated sugar was used as the donor and showed a high thermal tolerance with the most favorable enzymatic activity at 100 degrees C. In addition, we also found that the alpha-amylases and CGTase, that is, GH13 family enzymes, all exhibited similar activities, which were able to catalyze glucosylation in steroidal saponins. But other kinds of amylases, such as gamma-amylase (GH15 family), had no such activity under the same reaction conditions.
Zhou
Wen-bin
WB
Beijing Institute of Radiation Medicine, No. 27 Tai-ping Road, Haidian District, Beijing 100850, People's Republic of China.
Feng
Bing
B
Huang
Hong-zhi
HZ
Qin
Yu-juan
YJ
Wang
Yong-ze
YZ
Kang
Li-ping
LP
Zhao
Yang
Y
Wang
Xiao-nan
XN
Cai
Yun
Y
Tan
Da-wei
DW
Ma
Bai-ping
BP
D055162:Biocatalysis; D002240:Carbohydrate Sequence; D004789:Enzyme Activation; D005964:Glucosyltransferases / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D008968:Molecular Conformation; D012503:Saponins / Q000737:chemistry / Q000378:metabolism*; D013237:Stereoisomerism; D013256:Steroids / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors
D012503:Saponins; D013256:Steroids; C523027:timosaponin B-II; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase