S138: A strategy for the production of drug metabolites based on bacterial cytochrome P450s—Challenges and solutions

Microbial models for human drug metabolism have been an effective method to produce metabolites in quantities suitable for testing and are more attractive than other methods such as chemical synthesis.  In both microbes and mammals, cytochrome P450 enzyme activity is responsible for drug metabolism.  The bacterium Actinoplanes sp. ATCC 53771 has been found to have similar drug metabolism profile for several drugs as humans.  We have one cytochrome P450, CYP107E4, which is able to bind a variety of drug targets including diclofenac, rapamycin, tacrolimus and cyclosporin.  Furthermore, we have been able to produce metabolites of diclofenac in vitro. Utilizing E. coli as a host for metabolite production has the advantage of containing no native P450 systems that would inhibit or compete with the desired drug metabolism, but this also presents several challenges.  We have shown that coexpression of the hemA gene from R. sphaeroides supports efficient expression of the active form of CYP107E4 without supplementation of an expensive heme precursor.  Another challenge towards metabolite production using E. coli is the lack of efficient redox partners.  Bacterial electron transport systems from characterized bacteria such as Streptomyces have potential to support CYP107E4 turnover, or identification of the native redox partners from Actinoplanes sp. may be possible.  Although there is work remaining to be done, we have shown that there is potential for engineering an efficient bacterial system for drug metabolite production.