Mutability landscape guided engineering of a promiscuous microbial glycosyltransferase for regioselective synthesis of salidroside and icariside D2.
Int J Biol Macromol
; 263(Pt 1): 130229, 2024 Apr.
Article
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| MEDLINE
| ID: mdl-38378110
ABSTRACT
Microbial glycosyltransferases efficiently synthesize glucosides and have garnered increasing interest. However, limited regioselectivity has impeded their broad application, particularly in the pharmaceutical industry. In this study, the UDP-glycosyltransferase YjiC from Bacillus licheniformis (BlYjiC) was engineered to achieve the bidirectional regioselective glycosylation of tyrosol and its derivatives. Initially, site-directed saturation mutagenesis was performed on two newly identified substrate-binding cavities in the acceptor pocket of BlYjiC to provide a comprehensive blueprint of the interplay between mutations and function (mutability landscape). Iterative saturation mutagenesis was performed, guided by the mutability landscape. Two highly regioselective mutants M6 (M112L/I325Y/L70R/Q136E/I67E/M77R) and M2' (M112D/I62L) were generated, exhibiting >99 % regioselectivity toward the alcoholic and phenolic hydroxyl of tyrosol, respectively, compared with the wild-type (product mixture 5149 %). Both mutants exhibited excellent regioselectivity toward several dihydroxy phenolic substrates, offering valuable biocatalysts for the regioselective synthesis of glucosides. Their application was confirmed in a short synthesis of salidroside (3.6 g/L) and icariside D2 (2.4 g/L), which exhibited near-perfect regioselectivity. This study provides valuable insights into future protein engineering of similar enzymes and opens new avenues for their practical applications.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Alcohol Feniletílico
/
Fenoles
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Glicosiltransferasas
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Glucósidos
Idioma:
En
Revista:
Int J Biol Macromol
/
Int. j. biol. macromol
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International journal of biological macromolecules
Año:
2024
Tipo del documento:
Article
País de afiliación:
China