Characterization of a α-l-rhamnosidase from Bacteroides thetaiotaomicron with high catalytic efficiency of epimedin C.
Bioorg Chem
; 81: 461-467, 2018 12.
Article
em En
| MEDLINE
| ID: mdl-30243237
In this study, a α-l-rhamnosidase gene from Bacteroides thetaiotaomicron VPI-5482 was cloned and expressed in Escherichia coli. The specific activity of rhamnosidase was 0.57 U/mg in LB medium with 0.1â¯mM Isopropyl ß-d-Thiogalactoside (IPTG) induction at 28⯰C for 8â¯h. The protein was purified by Ni-NTA affinity, which molecular weight approximately 83.3â¯kDa. The characterization of BtRha was determined. The optimal activity was at 55⯰C and pH 6.5. The enzyme was stable in the pH range 5.0-8.0 for 4â¯h over 60%, and had a 1-h half-life at 50⯰C. The Kcat and Km for p-nitrophenyl-α-l-rhamnopyranoside (pNPR) were 1743.29â¯s-1 and 2.87â¯mM, respectively. The α-l-rhamnosidase exhibited high selectivity to cleave the α-1,2 and α-1,6 glycosidic bond between rhamnoside and rhamnoside, rhamnoside and glycoside, respectively, which could hydrolyze rutin, hesperidin, epimedin C and 2â³-O-rhamnosyl icariside II. Under the optimal conditions, BtRha transformed epimedin C (1â¯g/L) to icariin by 90.5% in 4â¯h. This study provides the first demonstration that the α-l-rhamnosidase could hydrolyze α-1,2 glycosidic bond between rhamnoside and rhamnoside.
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MEDLINE
Assunto principal:
Flavonoides
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Bacteroides thetaiotaomicron
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Glicosídeo Hidrolases
Idioma:
En
Ano de publicação:
2018
Tipo de documento:
Article