Variants of Flavin-containing Monooxygenase 3 (FMO3) Found in Subjects in an Updated Database of Genome Resources .

Single-nucleotide substitutions of human flavin-containing monooxygenase 3 ( FMO3 ) identified in the whole-genome sequences of the updated Japanese population reference panel (now containing 38,000 subjects) were investigated. In this study, 2 stop codon mutations, 2 frameshifts, and 43 amino-acid-substituted FMO3 variants were identified. Among these 47 variants, 1 stop codon mutation, 1 frameshift, and 24 substituted variants were already recorded in the National Center for Biotechnology Information database. Functionally impaired FMO3 variants are known to be associated with the metabolic disorder trimethylaminuria; consequently, the enzymatic activities of the 43 substituted FMO3 variants were investigated. Twenty-seven recombinant FMO3 variants expressed in bacterial membranes had similar activities toward trimethylamine N -oxygenation (~75-125%) to that of wild-type FMO3 (98 min -1 ). However, 6 recombinant FMO3 variants (Arg51Gly, Val283Ala, Asp286His, Val382Ala, Arg387His, and Phe451Leu) had moderately decreased (≈50%) activities toward trimethylamine N -oxygenation, and 10 recombinant FMO3 variants (Gly11Asp, Gly39Val, Met66Lys, Asn80Lys, Val151Glu, Gly193Arg, Arg387Cys, Thr453Pro, Leu457Trp, and Met497Arg) showed severely decreased FMO3 catalytic activity (<10%). Because of the known deleterious effects of FMO3 C- terminal stop codons, the four truncated FMO3 variants (Val187SerfsTer25, Arg238Ter, Lys416SerfsTer72, and Gln427Ter) were suspected to be inactive with respect to trimethylamine N- oxygenation. The FMO3 p.Gly11Asp and p.Gly193Arg variants were located within the conserved sequences of flavin adenine dinucleotide (positions 9-14) and NADPH (positions 191-196) binding sites, which are important for FMO3 catalytic function. Whole-genome sequence data and kinetic analyses revealed that 20 of the 47 nonsense or missense FMO3 variants had moderately or severely impaired activity toward N- oxygenation of trimethylaminuria. Significance Statement The number of single-nucleotide substitutions in human FMO3 recorded in the expanded Japanese population reference panel database was updated. One novel stop mutation, FMO3 p.Gln427Ter ; one novel frameshift (p.Lys416SerfsTer72); and 19 novel amino-acid-substituted FMO3 variants were identified, along with p.Arg238Ter, p.Val187SerfsTer25, and 24 amino-acid-substituted variants already recorded with individual rs numbers. Recombinant FMO3 Gly11Asp, Gly39Val, Met66Lys, Asn80Lys, Val151Glu, Gly193Arg, Arg387Cys, Thr453Pro, Leu457Trp, and Met497Arg variants showed severely decreased FMO3 catalytic activity, possibly associated with the metabolic disorder trimethylaminuria.