Open AccessHeterologous coexpression of the benzoate‐para‐hydroxylase CYP53B1 with different cytochrome P450 reductases in various yeasts
Author(s) -
Theron Chrispian W.,
Labuschagné Michel,
Albertyn Jacobus,
Smit Martha S.
Publication year - 2019
Publication title -
microbial biotechnology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.287
H-Index - 74
ISSN - 1751-7915
DOI - 10.1111/1751-7915.13321
Subject(s) - yarrowia , yeast , biology , cytochrome p450 , kluyveromyces marxianus , heterologous expression , reductase , saccharomyces cerevisiae , heterologous , monooxygenase , biochemistry , enzyme , recombinant dna , gene
Summary Cytochrome P450 monooxygenases (P450) are enzymes with high potential as biocatalysts for industrial applications. Their large‐scale applications are, however, limited by instability and requirement for coproteins and/or expensive cofactors. These problems are largely overcome when whole cells are used as biocatalysts. We previously screened various yeast species heterologously expressing self‐sufficient P450s for their potential as whole‐cell biocatalysts. Most P450s are, however, not self‐sufficient and consist of two or three protein component systems. Therefore, in the present study, we screened different yeast species for coexpression of P450 and P450‐reductase (CPR) partners, using CYP53B1 from Rhodotorula minuta as an exemplary P450. The abilities of three different coexpressed CPR partners to support P450 activity were investigated, two from basidiomycetous origin and one from an ascomycete. The various P450‐CPR combinations were cloned into strains of Saccharomyces cerevisiae, Kluyveromyces marxianus, Hansenula polymorpha, Yarrowia lipolytica and Arxula adeninivorans , using a broad‐range yeast expression vector. The results obtained supported the previous finding that recombinant A. adeninivorans strains perform excellently as whole‐cell biocatalysts. This study also demonstrated for the first time the P450 reductase activity of the CPRs from R. minuta and U. maydis . A very interesting observation was the variation in the supportive activity provided by the different reductase partners tested and demonstrated better P450 activity enhancement by a heterologous CPR compared to its natural partner CPR. This study highlights reductase selection as a critical variable for consideration in the pursuit of optimal P450‐based catalytic systems. The usefulness of A. adeninivorans as both a host for recombinant P450s and whole‐cell biocatalyst was emphasized, supporting earlier findings.