High-level expression and characterization of a stereoselective lipase from Aspergillus oryzae in Pichia pastoris.
Protein Expr Purif
; 155: 1-7, 2019 03.
Article
em En
| MEDLINE
| ID: mdl-30389593
ABSTRACT
Pichia pastoris expression is a mature and efficient eukaryotic expression system. In this work, Aspergillus oryzae lipase (AOL, with the molecular mass of 28â¯kDa), which can perform highly stereoselective hydrolysis of (R, S)-methyl 2-(4-hydroxyphenoxy) propanoate, was expressed in P. pastoris X-33. The specific activity of AOL was 432 U/mg, which was obtained by fed-batch cultivation in a 5â¯L bioreactor using a methanol feeding strategy. After fermentation, the supernatant was concentrated by ultrafiltration with a 10â¯kDa cut-off membrane and purified with DEAE-Sepharose™ FF ion-exchange chromatography and phenyl Seflnose™ 6 FF hydrophobic interaction chromatography. The purified lipase activity reached 5509 U/mg. AOL showed high activity toward short-chain triacylglyceride (C4), and the optimum temperature and pH were 40⯰C and 8.0, respectively. The purified enzyme activity was inhibited by Zn2+ and Cu2+. Moreover, the Km and Vmax values were 1â¯mM and 32.89â¯mmol/min, respectively.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Pichia
/
Aspergillus oryzae
/
Lipase
Idioma:
En
Revista:
Protein Expr Purif
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
China