Construction and characterization of a functional chimeric laccase from metagenomes suitable as a biocatalyst

AMB Express

Hiroshi Toda¹  Yuya Hayashi¹  Yuna Yamamoto¹  Daichi Tanaka¹  Nobuya Itoh¹  Serina Honda¹ 

[1] Biotechnology and Pharmaceutical Engineering Research Center and Department of Biotechnology, Toyama Prefectural University, 5180 Kurokawa, 939-0398, Imizu, Toyama, Japan;

关键词: Metagenome;    S-GAM technique;    Multi-copper oxidase (MCO);    Laccase;    Chimeric enzyme;    Biocatalyst;   

DOI  :  10.1186/s13568-021-01248-y

来源: Springer

【 摘 要 】

Screening of gene-specific amplicons from metagenomes (S-GAM) is an efficient technique for the isolation of homologous genes from metagenomes. Using the S-GAM approach, we targeted multi-copper oxidase (MCO) genes including laccase and bilirubin oxidase (BOX) in soil and compost metagenomes, and successfully isolated novel MCO core regions. These core enzyme genes shared approximately 70% identity with that of the putative MCO from Micromonospora sp. MP36. According to the principle of S-GAM, the N- and C-terminal regions of the deduced products of the mature gene come from the known parent gene, which should be homologous and compatible with the target gene. We constructed two different MCO hybrid genes using Bacillus subtilis BOX and Micromonospora sp. MP36 MCO, to give Bs-mg-mco and Mic-mg-mco, respectively. The constructed chimeric MCO genes were fused with the maltose-binding protein (MBP) gene at the N-terminus for expression in Escherichia coli cells. We found that MBP-Mic-mg-MCO/Mic-mg-MCO possessed the characteristic properties of laccase, although MBP-Bs-mg-MCO had no activity. This novel laccase (Mic-mg-MCO) demonstrated unique substrate specificity, sufficient activity at neutral pH, and high thermal stability, which are suitable properties for its use as a laccase biocatalyst.

【 授权许可】

CC BY