Unspecific peroxygenases(UPOs, EC 1.11.2.1) is a kind of thioheme enzyme capable of catalyzing various oxidations of inert C–H bonds using H_(2)O_(2) as an oxygen donor without cofactors. However, the enhancement of ...Unspecific peroxygenases(UPOs, EC 1.11.2.1) is a kind of thioheme enzyme capable of catalyzing various oxidations of inert C–H bonds using H_(2)O_(2) as an oxygen donor without cofactors. However, the enhancement of the H_(2)O_(2) tolerance of UPOs is always challenging. In this study, the A161C mutant of r Dca UPO,which originates from Daldinia caldariorum, was found to be highly H_(2)O_(2)-resistant. Compared with the wild type, the mutant r Dca UPO-A161C showed a 10-h prolonged half-life and a 64% improved enzyme activity when incubated in 10 mmol/L H_(2)O_(2). The crystal structure analysis at 1.47 A showed that r Dca UPOA161C exhibited 10 α-helixes(cyan) and a series of ordered rings, forming a single asymmetric spherical structure. The two conserved domains near heme formed an active site with the catalytic PCP and EHD regions(Glu86, His87, Asp88 residues). The H_(2)O_(2) tolerance of r Dca UPO-A161C was preliminarily explored by comparing its structure with the wild type. Notably, r Dca UPO-A161C showed significantly higher catalytic efficiency than the wild type for the production of hydroxyl fatty acids. This study is anticipated to provide an insight into the structure-function relationship and expand potential applications of UPOs.展开更多
基金supported by the National Natural Science Foundation of China (No.32001633)the Key Program of Natural Science Foundation of China (No.31930084)Guangzhou Science and technology planning project (No.202102020370)。
文摘Unspecific peroxygenases(UPOs, EC 1.11.2.1) is a kind of thioheme enzyme capable of catalyzing various oxidations of inert C–H bonds using H_(2)O_(2) as an oxygen donor without cofactors. However, the enhancement of the H_(2)O_(2) tolerance of UPOs is always challenging. In this study, the A161C mutant of r Dca UPO,which originates from Daldinia caldariorum, was found to be highly H_(2)O_(2)-resistant. Compared with the wild type, the mutant r Dca UPO-A161C showed a 10-h prolonged half-life and a 64% improved enzyme activity when incubated in 10 mmol/L H_(2)O_(2). The crystal structure analysis at 1.47 A showed that r Dca UPOA161C exhibited 10 α-helixes(cyan) and a series of ordered rings, forming a single asymmetric spherical structure. The two conserved domains near heme formed an active site with the catalytic PCP and EHD regions(Glu86, His87, Asp88 residues). The H_(2)O_(2) tolerance of r Dca UPO-A161C was preliminarily explored by comparing its structure with the wild type. Notably, r Dca UPO-A161C showed significantly higher catalytic efficiency than the wild type for the production of hydroxyl fatty acids. This study is anticipated to provide an insight into the structure-function relationship and expand potential applications of UPOs.
