Overexpression of chaperones GroEL/ES from Escherichia coli enhances the indigo biotransformation production of cytochrome P450 BM3 mutant

Abstract

Abstract The self-sufficient cytochrome P450 BM3 mutant (A74G/F87V/D168H/L188Q) could be used as the biocatalyst in the whole-cell catalysis process of indigo. Nevertheless, the bioconversion yield of indigo is low under normal cultivation conditions (37˚C, 250 rpm). In this paper, a recombinant E. coli BL21(DE3) strain was constructed to co-express both P450 BM3 mutant gene and GroEL/ES genes to investigate whether GroEL/ES could promote the indigo bioconversion production in E. coli. Results revealed that GroEL/ES system increased indigo bioconversion yield significantly, and the indigo bioconversion production of strain co-expressing P450 BM3 mutant and GroEL/ES was 21 times that of strain expressing P450 BM3 mutant alone. P450 BM3 enzyme content and in vitro indigo formation yield were determined to investigate the underlying reasons for the improved indigo bioconversion production. And it revealed that GroEL/ES did not increase indigo bioconversion production through improving P450 BM3 enzyme content and its enzymatic transformation efficiency. We also demonstrated that GroEL/ES could improve the intracellular nicotinamide adenine dinucleotide phosphate (NADPH)/NADP+ ratio. Given NADPH was an important coenzyme in the indigo catalytic process, the underlying mechanism of the increased indigo bioconversion yield was probably related to the improved intracellular NADPH/NADP+ ratio.