Title : Random mutagenesis and selection of organic solvent-stable haloperoxidase from Streptomyces aureofaciens - Yamada_2015_Biotechnol.Prog_31_917 |
Author(s) : Yamada R , Higo T , Yoshikawa C , China H , Yasuda M , Ogino H |
Ref : Biotechnol Prog , 31 :917 , 2015 |
Abstract :
Haloperoxidases are useful oxygenases involved in halogenation of a range of water-insoluble organic compounds and can be used without additional high-cost cofactors. In particular, organic solvent-stable haloperoxidases are desirable for enzymatic halogenations in the presence of organic solvents. In this study, we adopted a directed evolution approach by error-prone polymerase chain reaction to improve the organic solvent-stability of the homodimeric BPO-A1 haloperoxidase from Streptomyces aureofaciens. Among 1,000 mutant BPO-A1 haloperoxidases, an organic solvent-stable mutant OST48 with P123L and P241A mutations and a high active mutant OST959 with H53Y and G162R mutations were selected. The residual activity of mutant OST48 after incubation in 40% (v/v) 1-propanol for 1 h was 1.8-fold higher than that of wild-type BPO-A1. In addition, the OST48 mutant showed higher stability in methanol, ethanol, dimethyl sulfoxide, and N,N-dimethylformamide than wild-type BPO-A1 haloperoxidase. Moreover, after incubation at 80 degrees C for 1 h, the residual activity of mutant OST959 was 4.6-fold higher than that of wild-type BPO-A1. Based on the evaluation of single amino acid-substituted mutant models, stabilization of the hydrophobic core derived from P123L mutation and increased numbers of hydrogen bonds derived from G162R mutation led to higher organic solvent-stability and thermostability, respectively. |
PubMedSearch : Yamada_2015_Biotechnol.Prog_31_917 |
PubMedID: 26033926 |
Gene_locus related to this paper: strau-brpa1 |
Gene_locus | strau-brpa1 |
Yamada R, Higo T, Yoshikawa C, China H, Yasuda M, Ogino H (2015)
Random mutagenesis and selection of organic solvent-stable haloperoxidase from Streptomyces aureofaciens
Biotechnol Prog
31 :917
Yamada R, Higo T, Yoshikawa C, China H, Yasuda M, Ogino H (2015)
Biotechnol Prog
31 :917