A combined computational and experimental study on selective flucloxacillin hydroxylation by cytochrome P450 BM3 variants.

A combined computational and experimental study on selective flucloxacillin hydroxylation by cytochrome P450 BM3 variants. J Inorg Biochem. 2018 Apr 24;184:115-122 Authors: Luirink RA, Dekker SJ, Capoferri L, Janssen LFH, Kuiper CL, Ari ME, Vermeulen NPE, Vos JC, Commandeur JNM, Geerke DP Abstract The 5'-hydroxymethyl metabolite of the penicillin based antibiotic flucloxacillin (FLX) is considered to be involved in bile duct damage occurring in a small number of patients. Because 5'-hydroxymethyl FLX is difficult to obtain by organic synthesis, biosynthesis using highly active and regioselective biocatalysts would be an alternative approach. By screening an in-house library of Cytochrome P450 (CYP) BM3 mutants, mutant M11 L437E was identified as a regioselective enzyme with relatively high activity in production of 5'-hydroxymethyl FLX as was confirmed by mass spectrometry and NMR. In contrast, incubation of M11 L437E and other mutants with oxacillin (OX, which differs from FLX by a lack of aromatic halogens) resulted in formation of two metabolites. In addition to 5'-hydroxymethyl OX we identified a product resulting from aromatic hydroxylation. In silico studies of both FLX and OX with three CYP BM3 mutants revealed substrate binding poses allowing for 5'-methyl hydroxylation, as well as binding poses with the aromatic moiety in the vicinity of the heme iron for which the corresponding product of aromatic hydroxylation was not obse...
Source: Journal of Inorganic Biochemistry - Category: Biochemistry Authors: Tags: J Inorg Biochem Source Type: research