A novel cytochrome P450 mono‐oxygenase from Streptomyces platensis resembles activities of human drug metabolizing P450s

Abstract

Cytochrome P450 mono-oxygenases (P450) are versatile enzymes which play essential roles in C-source assimilation, secondary metabolism, and in degradations of endo- and exogenous xenobiotics. In humans, several P450 isoforms constitute the largest part of phase I metabolizing enzymes and catalyze oxidation reactions which convert lipophilic xenobiotics, including drugs, to more water soluble species. Recombinant human P450s and microorganisms are applied in the pharmaceutical industry for the synthesis of drug metabolites for pharmacokinetics and toxicity studies. Compared to the membrane-bound eukaryotic P450s, prokaryotic ones exhibit some advantageous features, such as high stability and generally easier heterologous expression. Here, we describe a novel P450 from Streptomyces platensis DSM 40041 classified as CYP107L that efficiently converts several commercial drugs of various size and properties. This P450 was identified by screening of actinobacterial strains for amodiaquine and ritonavir metabolizing activities, followed by genome sequencing and expression of the annotated S. platensis P450s in Escherichia coli. Performance of CYP107L in biotransformations of amodiaquine, ritonavir, amitriptyline, and thioridazine resembles activities of the main human metabolizing P450s, namely CYPs 3A4, 2C8, 2C19, and 2D6. For application in the pharmaceutical industry, an E. coli whole-cell biocatalyst expressing CYP107L was developed and evaluated for preparative amodiaquine metabolite production.

Accessing the Biocatalytic Potential for C−H‐Activation by Targeted Genome Mining and Screening

“…M8 with an m/z ratio of 735.3 is possibly corresponding to an oxidation of metabolite M5 to a ketone. This metabolite was described before in a biotransformation of ritonavir with a P450 of the strain Streptomyces platensis . Furthermore, products with the m/z ratio of 606.3 and 622.3 could be identified which were described as degradation products of ritonavir before and were also present in the sample directly taken after substrate addition .…”

“…The screening resulted in nine strains showing good bioconversion rates of cyclosporine into the two human hydroxylated (AM1 and AM9) and N ‐demethylated (AM4) derivatives . In a recent study a novel P450 from the actinomycete Streptomyces platensis was identified showing high activity for the conversion of several pharmaceutical compounds while producing equivalent metabolites to the human metabolizing P450s 3A4, 2C8, 2C19, and 2D6 …”

Two‐step Screening for Identification of Drug‐metabolizing Bacterial Cytochromes P450 with Diversified Selectivity

“…According to the online P450 database CYPED 31 genes belong to the CYP105 and the CYP107 families. Fourteen of those genes (13 of S. platensis and 1 of P. autotrophica ) have been investigated in previous studies . One cyp gene in P. autotrophica has a length of only 378 bp and was not considered further.…”

“…Recently, our group described nine metabolites of ritonavir biotransformed by the parent organism of both P450s Streptomyces platensis DSM 40041. Metabolites were analyzed by LC/MS and LC/MS‐MS analysis . The observed mass differences and retention times for 1 b – d correspond to the N‐dealkylated and N‐demethylated metabolites described there (Table S7).…”

“…In this study we combined the advantages of both screening methods aiming at identification and characterization of drug‐metabolizing P450 enzymes from Streptomyces platensis DSM 40041 and Pseudonocardia autotrophica DSM 535. Both strains are utilized by pharmaceutical companies for the oxidation of various drugs . Using substrate profiling with three model drugs namely ritonavir, testosterone and amitriptyline in the first step, two P450s from S. platensis with broad substrate spectra and complementary selectivities were selected.…”