Allerston, C.K.J.; (2008) Flavin-containing monooxygenase 3 (FM03): Evolution, protein structure and causative mutations of Trimethylaminuria. Doctoral thesis , University of London.

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Abstract

Flavin-containing monooxygenase 3 (FM03) is a hepatic, microsomal enzyme important in the Phase I metabolism of xenobiotics. Numerous single nucleotide polymorphisms (SNPs) in FM03 have been identified and shown to affect the catalytic activity of the enzyme, with some destroying the activity altogether, causing the distressing disorder, Trimethyalminuria (TMAU), in humans. Genotyping was performed at the FM03 locus on several individuals displaying symptoms of TMAU. Two new FM03 variants, R238Q and R492Q, were discovered in this investigation, with kinetic studies suggesting that R238Q destroys FM03 catalysis. Kinetic studies of FM03 common polymorphisms showed that the E158K/E308G displayed significantly reduced catalytic activity compared with either single variant alone. E158K/V257M also displayed significantly reduced catalytic activity, but not compared to either single variant alone. The pharmacogenetic implications are discussed with particular reference to the recent finding that E158K/E308G has been associated with reduced polyp formation in patients with familial adenomatous polyposis who were treated with sulindac sulphide, an FM03 substrate. The evolutionary history of FM03 was probed using sequence data of genomic DNA in a Japanese cohort of potential TMAU sufferers and control individuals. Mutational relationships among haplotypes were inferred and time depth of the variation and ages of individual mutations were estimated by coalescent analysis, with test statistics used to detect departure from neutral evolution. A case of balancing selection is proposed at the FM03 locus. In an attempt to understand the structural and biophysical consequences of FM03 variants, a homology model of FM03 was generated, refined and validated. Flavin adenine dinucleotide (FAD), an FM03 cofactor, was also modeled into the FM03 model and the interactions between the enzyme and cofactor predicted.

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