Directed Evolution of a Cytochrome P450 Monooxygenase for Alkane Oxidation

“…TB medium is the most frequently reported medium for its use in the production of recombinant P450 enzymes in E. coli [15,16]. In this study, experiments with TB medium for the production of P450SMO were also studied under the optimal flask fermentation conditions.…”

Section: Resultsmentioning

confidence: 91%

Improved expression of recombinant cytochrome P450 monooxygenase in Escherichia coli for asymmetric oxidation of sulfides

Zhang

2010

Bioprocess Biosyst Eng

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Escherichia coli BL21 as production strain for the production of cytochrome P450 monooxygenase (P450SMO) from Rhodococcus sp. in high yields was developed. The expression was first optimized with a series of flask experiments testing several key parameters for their influence on the expression level and enzyme activity. The optimal process parameters found in the flask experiments were verified in a cultivation process in a 5-L bioreactor. Glycerol proved to be superior over glucose as carbon source. Low dissolved oxygen (DO) concentration (<10%) during expression was found to be critical for active P450s production, resulting in expression level of 400 nM for P450SMO. Intact cells were used to establish an efficient bioconversion system for the production of sulfoxidation product. With p-chlorothioanisole as a representative substrate, the desired product (S-sulfoxide) was afforded with 99% ee and highest production of 130 mg/L within 12 h.

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“…The CO difference spectrum experiments exhibited a formation of a peak at 450 nm indicating the successful expression of the active enzyme in the heterogeneous host. The formation of a maximum peak at 450 nm during the reduction of CO was formerly described for CYP450 (Farinas et al 2001).…”

Section: Identification Of a Novel Cyclohexane Monooxygenasementioning

confidence: 99%

Novel cyclohexane monooxygenase from Acidovorax sp. CHX100

Salamanca

Karande

Schmid

et al. 2015

Appl Microbiol Biotechnol

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Acidovorax sp. CHX100 has a remarkable ability for growth on short cycloalkanes (C5-C8) as a sole source of carbon and energy under aerobic conditions via an uncharacterized mechanism. Transposon mutagenesis of Acidovorax sp. CHX100 revealed a novel cytochrome P450 monooxygenase (CYP450chx) which catalyzed the transformation of cyclohexane to cyclohexanol. Primer walking methods categorized CYP450chx as cytochrome P450 class I taking into account its operon structure: monooxygenase, FAD oxidoreductase, and ferredoxin. CYP450chx was successfully cloned and expressed in Escherichia coli JM109. The activity of CYP450chx was demonstrated by means of the indole co-oxidation. Biotransformation capability of CYP450chx was confirmed through the catalysis of cycloalkanes (C5-C8) to their respective cyclic alcohols.

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Directed Evolution of a Cytochrome P450 Monooxygenase for Alkane Oxidation

Cited by 33 publications

References 0 publications

Alkene epoxidation catalyzed by cytochrome P450 BM-3 139-3

Alkene epoxidation catalyzed by cytochrome P450 BM-3 139-3

Improved expression of recombinant cytochrome P450 monooxygenase in Escherichia coli for asymmetric oxidation of sulfides

Novel cyclohexane monooxygenase from Acidovorax sp. CHX100

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