A panel of cytochrome P450 BM3 variants to produce drug metabolites and diversify lead compounds.
Herein we demonstrate that a small panel of variants of cytochrome P450 BM3 from Bacillus megaterium covers the breadth of reactivity of human P450s by producing 12 of 13 mammalian metabolites for two marketed drugs, verapamil and astemizole, and one research compound. The most active enzymes support preparation of individual metabolites for preclinical bioactivity and toxicology evaluations. Underscoring their potential utility in drug lead diversification, engineered P450 BM3 variants also produce novel metabolites by catalyzing reactions at carbon centers beyond those targeted by animal and human P450s. Production of a specific metabolite can be improved by directed evolution of the enzyme catalyst. Some variants are more active on the more hydrophobic parent drug than on its metabolites, which limits production of multiply-hydroxylated species, a preference that appears to depend on the evolutionary history of the P450 variant.
Division of Chemistry and Chemical Engineering 210-41, California Institute of Technology, Pasadena, CA 91125-4100, USA.
Andrew M
AM
Sawayama
Michael M Y
MM
Chen
Palaniappan
P
Kulanthaivel
Ming-Shang
MS
Kuo
Horst
H
Hemmerle
Frances H
FH
Arnold
D016589:Astemizole / Q000737:chemistry / Q000378:metabolism; D001410:Bacillus megaterium / Q000201:enzymology; D055162:Biocatalysis; D020134:Catalytic Domain; D002867:Chromones / Q000737:chemistry / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D019143:Evolution, Molecular; D006801:Humans; D009025:Morpholines / Q000737:chemistry / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014700:Verapamil / Q000737:chemistry / Q000378:metabolism
D002867:Chromones; D009025:Morpholines; D004364:Pharmaceutical Preparations; D011993:Recombinant Fusion Proteins; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D016589:Astemizole; D003577:Cytochrome P-450 Enzyme System; D014700:Verapamil