Cytochromes P450 as promising catalysts for biotechnological application: chances and limitations

Abstract

Cytochromes P450 (CYPs) belong to the superfamily of heme b containing monooxygenases with currently more than 21,000 members. These enzymes accept a vast range of organic molecules and catalyze diverse reactions. These extraordinary capabilities of CYP systems that are unmet by other enzymes make them attractive for biotechnology. However, the complexity of these systems due to the need of electron transfer from nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) via redox partner proteins for the initial hydroxylation step limits a broader technical implementation of CYP enzymes. There have been several reviews during the past years tackling the potential CYPs for synthetic application. The aim of this review is to give a critical overview about possibilities and chances for application of these interesting catalysts as well as to discuss drawbacks and problems related to their use. Solutions to overcome these limitations will be demonstrated, and several selected examples of successful CYP applications under industrial conditions will be reviewed.

CYP267A1 and CYP267B1 from Sorangium cellulosum So ce56 are Highly Versatile Drug Metabolizers

“…However, an important bottleneck in the application of P450 enzymes in biotechnological processes is often the efficiency of the redox system (Bernhardt and Urlacher, 2014). Therefore, we first identified efficient autologous redox partners to transfer electrons to the CYP267 family.…”

“…The established CYP267-Fdx8-FdR_B whole-cell systems are an excellent starting point for further optimizations in view of biotechnological upscaling and optimization (Bernhardt and Urlacher, 2014). Optimizations such as changing expression and reaction conditions or engineering P450 enzymes could be potential topics of interest.…”

“…However, because of the release of side products and the difficulty on handling the microbes during such biotransformation process, there is now a great interest in cytochrome P450 (P450) enzymes for the production of drug metabolites. In general, P450 enzymes are versatile, heme-containing enzymes, which catalyze a variety of reactions highlighting them as essential candidates for biotechnological and pharmaceutical research (Bernhardt and Urlacher, 2014). It has been shown that the utilization of human P450 enzymes enables the sufficient production of human drug metabolites employing baculovirus-infected insect cells expressing CYP3A4 or CYP2D9 (Rushmore et al, 2000), fission yeast expressing CYP2D6 (Peters et al, 2007) or CYP2C9 (Drǎgan et al, 2011;Neunzig et al, 2012), and Escherichia coli cells expressing CYP3A4, CYP2C9, and CYP1A2 (Vail et al, 2005).…”

A Peroxygenase from Chaetomium globosum Catalyzes the Selective Oxygenation of Testosterone

“…In this context, enzymatic steroid hydroxylation catalyzed by engineered P450 mutants (e.g., of BM3) has gained attention for the preparation of diverse and unique hydroxylated steroids with pharmacological activity 8. ‐9 However, the use of isolated P450 for larger‐scale application is hampered by low activity, catalytic efficiency, and stability outside cells, the requirement for NAD(P)H (as electron donating co‐substrate), and the necessity for electron‐transferring partners 10…”

Regioselective Biocatalytic Hydroxylation of Fatty Acids by Cytochrome P450s

“…Challenges to overcome are the poor substrate solubility, low coupling efficiencies and the formation of side products due to over-oxidation or multiple oxidations [117]. Careful balance of the available (dissolved) oxygen in the system was demonstrated to be crucial for selective single-oxidations with an E. coli designer cell expressing BM3 and a pGEc47-fatty acid uptake system.…”