Flavin-containing Monooxygenase 3 Gene Research Articles

Regulation of flavin-containing mono-oxygenase (Fmo3) gene expression by steroids in mice and humans.

Abstract Flavin-containing mono-oxygenases (FMOs) are a family of microsomal chemical- and drug-metabolizing enzymes. FMO3 is a major FMO form in adult mouse and human liver. FMO3 mutations have been associated with the incidence and severity of trimethylaminuria (TMAU), a metabolic disorder characterized by the inability of the affected individual to metabolize the odorous trimethylamine to its non-odorous N-oxide. In addition to this primary genetic form, there are other forms of TMAU that support the hypothesis that FMO3 activity may be modulated by steroid hormones. To understand the molecular mechanism involved in the regulation of Fmo3 gene expression by steroid hormones, we performed this study in an in vitro cellular system, mouse liver cells, and on the human FMO3 gene. Dexamethasone, 5α-dihydrotestosterone, thyroid hormone, and progesterone had no effect on the accumulation of Fmo3 mRNA. The use of increased concentration of theophylline inhibited estrogen receptor α (ERα)-mediated transcription of Fmo3 mRNA. 17β-Estradiol inhibited Fmo3 mRNA accumulation. The use of ICI 164,384 abolished the inhibitory effect induced by estrogen. Gel-shift analyses showed a binding in the 5' region of the Fmo3 gene. This binding was abrogated by an excess of a cDNA containing an estrogen-responsive element. An estrogen-binding site was also present in the first intron of the human gene, as demonstrated by the gel-shift assay. Supershift experiments confirmed the binding of ERα in both mouse and human samples. Furthermore, chromatin immunoprecipitation assay confirmed the binding of ERα in the promoter region of mouse Fmo3 and in the first intron of the human FMO3 gene. Thus, 17β-estradiol plays a fundamental role in the regulation of Fmo3 gene transcription.

Potential for drug interactions mediated by polymorphic flavin-containing monooxygenase 3 in human livers

Human flavin-containing monooxygenase 3 (FMO3) in the liver catalyzes a variety of oxygenations of nitrogen- and sulfur-containing medicines and xenobiotic substances. Because of growing interest in drug interactions mediated by polymorphic FMO3, benzydamine N-oxygenation by human FMO3 was investigated as a model reaction. Among the 41 compounds tested, trimethylamine, methimazole, itopride, and tozasertib (50 μM) suppressed benzydamine N-oxygenation at a substrate concentration of 50 μM by approximately 50% after co-incubation. Suppression of N-oxygenation of benzydamine, trimethylamine, itopride, and tozasertib and S-oxygenation of methimazole and sulindac sulfide after co-incubation with the other five of these six substrates was compared using FMO3 proteins recombinantly expressed in bacterial membranes. Apparent competitive inhibition by methimazole (0–50 μM) of sulindac sulfide S-oxygenation was observed with FMO3 proteins. Sulindac sulfide S-oxygenation activity of Arg205Cys variant FMO3 protein was likely to be suppressed more by methimazole than wild-type or Val257Met variant FMO3 protein was. These results suggest that genetic polymorphism in the human FMO3 gene may lead to changes of drug interactions for N- or S-oxygenations of xenobiotics and endogenous substances and that a probe battery system of benzydamine N-oxygenation and sulindac sulfide S-oxygenation activities is recommended to clarify the drug interactions mediated by FMO3.

Relationships between flavin‐containing mono‐oxygenase 3 (FMO3) genotype and trimethylaminuria phenotype in a Japanese population

The aim of this study was to investigate relationships between flavin-containing mono-oxygenase 3 (FMO3) genotype and phenotype (conversion of odorous trimethylamine into non-odorous trimethylamine N-oxide) in a large Japanese cohort suffering from trimethylaminuria. Urinary excretion of trimethylamine and trimethylamine N-oxide was determined for 102 volunteers with self-reporting symptoms of trimethylaminuria. For each we determined the sequence of the entire coding region, plus 1.3 kb of flanking intronic and 2.5 kb of the upstream region of the FMO3 gene. The affect of upstream variants on transcription was determined with a reporter gene assay. Seventy-eight subjects were diagnosed as suffering from trimethylaminuria, based on urinary excretion of <90% of total TMA as TMA N-oxide. Of these, 13 were classified as severe, 56 as moderate and nine as mild cases, excreting <43%, 48-70% and 73-83% of trimethylamine as trimethylamine N-oxide, respectively. Twenty-seven mutations were identified in FMO3, 15 in the coding region, of which eight abolish or severely impair FMO3 activity (Pro70Leu, Cys197fsX, Thr201Lys, Arg205Cys, Met260Val, Trp388Ter, Gln470Ter and Arg500Ter), and 12 in the upstream region. The mutations segregate into 19 haplotypes, including four different combinations of upstream mutations, each of which reduces transcriptional activity in comparison with the ancestral upstream sequence of FMO3. Comparisons of genotype and phenotype reveal that severe trimethylaminuria is caused by loss of function mutations in FMO3. For moderate and mild cases the situation is more complex, with most resulting from factors other than FMO3 genotype. Our results have implications for the diagnosis and management of the disorder.

G476(P) Trimethylaminuria: A case report

BackgroundTrimethylaminura; also known as Fish-Odour Syndrome; is an inherited disorder characterised by a body odour likened to that of rotting fish. This metabolic condition caused by deficient N-oxidation and increased...

Quail FMO3 Gene Cloning, Tissue Expression Profiling, Polymorphism Detection and Association Analysis with Fishy Taint in Eggs

Quail eggs comprise a significant and favourable part of table eggs in certain countries. Some quail eggs, however, present fishy off-flavor which directly influences their quality. It is reported that flavin-containing monooxygenase 3 (FMO3) is associated with fish-odour trait in human and animal products. FMO3 is responsible for the degradation of trimethylamine (TMA) in vivo. Loss-of-function mutations in FMO3 gene can result in defective TMA N-oxygenation, giving rise to disorder known as “fish-odour syndrome” in human, as well as the fishy off-flavor in cow milk and chicken eggs. In order to reveal the genetic factor of fishy taint in quail eggs, we cloned the cDNA sequence of quail FMO3 gene, investigated FMO3 mRNA expression level in various tissues, detected SNPs in the coding region of the gene and conducted association analysis between a mutation and the TMA content in quail egg yolks. The 1888 bp cDNA sequence of quail FMO3 gene encoding 532 amino acids was obtained and characterized. The phylogenetic analysis revealed quail FMO3 had a closer relationship with chicken FMO3. The FMO3 mRNA was highly expressed in liver and kidney of quail. Nine SNPs were detected in the coding sequence of quail FMO3 gene, including a nonsense mutation (Q319X) which was significantly associated with the elevated TMA content in quail egg yolks. Genotype TT at Q319X mutation loci was sensitive to choline. With addition of choline in the feed, the quails with homozygote TT at the Q319X mutation loci laid fish-odour eggs, indicating an interaction between genotype and diet. The results indicated that Q319X mutation was associated with the fishy off-flavor in quail eggs. Identification of the unfavorable allele T of quail FMO3 gene can be applied in future quail breeding to eliminate fishy off-flavor trait in quail eggs.

Trimethylaminuria (fish odor syndrome): Genotype characterization among Portuguese patients

Trimethylaminuria (TMAu) or “fish odor syndrome” is a metabolic disorder characterized by the inability to convert malodorous dietarily-derived trimethylamine (TMA) to odorless TMA N-oxide by the flavin-containing monooxygenase 3 (FMO3). Affected individuals unable to complete this reaction exude a “fishy” body odor due to the secretion of TMA in their corporal fluids leading to a variety of psychosocial problems. Interindividual variability in the expression of FMO3 gene may affect drug and foreign chemical metabolism in the liver and other tissues. Therefore, it is important to screen for common TMAu mutations but also extend the search to other genetic variants in order to correlate genotype and disease-associated phenotypes.In this study, 25 Portuguese patients with phenotype suggestive of TMAu were evaluated for molecular screening of the FMO3 gene. Herein, we found 16 variants in the FMO3 coding region, some of which had not been previously documented (Gly38Trp, Asp232Val, Thr307Pro, Ser310Leu). Whenever common variants (Glu158Lys, Glu308Gly) were considered in combination a distinct pattern between the control population and patients was observed, mainly in what concerns the presence of Lys158 and Gly308 in homozygous state.Further studies are necessary to clarify the pathogenicity of novel variants identified in this study, as well as the effect of the common single nucleotide polymorphisms, which may play an important role in disease presentation and/or protective mechanism to xenobiotics drugs or environment.

Molecular cloning, sequence characterization, SNP detection, and tissue expression analysis of duck FMO3 gene

Flavin-containing monooxygenase 3 (FMO3) is an important monooxygenase for catalytic oxygenation of many harmful xenobiotics. Mutations in the FMO3 gene have been identified as causing trimethylaminuria in human and fishy off-flavor in cow milk and chicken eggs. In this study, the full-length cDNA sequence of Pekin duck FMO3 gene was cloned, sequenced, and characterized. The full-length cDNA sequence consisted of 1,846 bp and contained a 1,599 bp open-reading frame encoding 532 amino acids. Duck FMO3 gene shared a similar nine exon-eight intron structure with chicken and human. The duck FMO3 putative protein sequence showed high identity with that of chicken (82 %), and relative low identity with those of mammals (61-66 %). We also found that the duck FMO3 gene was dramatically expressed in liver, lung, and kidney compared to that in other tissues in the ducks, indicating the possible roles the FMO3 gene could play in the three tissues. By bidirectional sequencing, we also found one nonsense mutation, 5 nonsynonymous, and 21 synonymous mutations in the coding region of the FMO3 gene in 11 duck breeds and some of them were predicted to be potentially associated with the activities of FMO3 protein.

Molecular and functional characterization of flavin-containing monooxygenases in cynomolgus macaque

Flavin-containing monooxygenases (FMOs), drug-metabolizing enzymes essential for the metabolism of endogenous biochemicals and foreign compounds, have been characterized in human (including FMO1-5 and FMO6P), but remain to be investigated in cynomolgus macaque. In this study, cDNAs of cynomolgus FMO1-5 and FMO6 were isolated and characterized. Amino acid sequences of cynomolgus FMO1-5, respectively, shared high sequence identities (94–98%) and were closely clustered in a phylogenetic tree, with human FMO1-5. Eight different transcripts, due to alternative splicing, were isolated for cynomolgus FMO6, which is highly identical (~96%) to human FMO6P. Among the 10 tissue types analyzed, cynomolgus FMO1, FMO2, FMO4, and FMO6 were most abundantly expressed in kidney, while cynomolgus FMO3 and FMO5 were most abundantly expressed in liver. In kidney and liver, the most abundantly expressed cynomolgus FMO genes were FMO1 and FMO3, respectively. Cynomolgus FMO1, FMO2, FMO3, and FMO5 metabolized benzydamine, and FMO1/FMO3 and FMO3 also metabolized methimazole and trimethylamine, respectively. Rates of benzydamine N-oxygenation (catalyzed by FMO3) varied (approximately 20-fold) among the 28 cynomolgus livers and were significantly correlated with FMO3 protein expression, indicating that the inter-animal variations in benzydamine N-oxygenation might be partly accounted for by the variable FMO3 expression. Cynomolgus FMO6 metabolized benzydamine only slightly, but minimal expression of FMO6 in all tissue precludes the importance of FMO6 in drug metabolism, unlike cynomolgus FMO1, FMO2, FMO3, and FMO5 which were all functional. Abundant expression of FMO1 and FMO3 in kidney and liver, respectively, suggest their importance in drug metabolism in cynomolgus macaque, similar to human.

Survey of variants of human flavin-containing monooxygenase 3 (FMO3) and their drug oxidation activities

Human flavin-containing monooxygenase 3 (FMO3, EC 1.14.13.8) in liver catalyzes a variety of oxygenations of nitrogen- and sulfur-containing medicines and xenobiotic substances. Loss-of-function mutations of the FMO3 gene, the enzyme responsible for trimethylamine N-oxygenation, cause the inherited disorder trimethylaminuria (also known as fish odor syndrome). In this mini-review, mutations of the FMO3 gene reported in the literature and in the National Center for Biotechnology Information single nucleotide polymorphism database were surveyed. Then, the activities of FMO3 variants in human liver microsomes and the activities of recombinantly expressed FMO3 variant proteins with respect to the oxygenation of nitrogen- and sulfur-containing drugs were summarized and the potential for drug interactions was demonstrated. Individual differences in FMO3 function were seen in subjects genotyped for homozygous FMO3 variants. Specific regions of the FMO3 C-terminus are required for functional activity. Naturally truncated FMO3 is believed to have barely detectable function, thereby explaining the relationship with severe impaired phenotypes. The present article provides fundamental, up-to-date information on the importance of human FMO3 in individual xenobiotic oxygenations, including those of new medicines and dietary-derived trimethylamine.

Regulation of FMO and PON Detoxication Systems in ALS Human Tissues

Amyotrophic lateral sclerosis (ALS) is an adult-onset, progressive, and fatal neurodegenerative disease with unknown etiology. Recent evidence suggests an association between the exposure to toxic environmental factors and sporadic ALS. The flavin-containing monooxygenases (FMOs) and paraoxonase (PONs) genes encode enzymes involved in xenobiotic detoxication and are associated with ALS. FMO and PON gene expression has been examined in the human central nervous system including human brain subregions defined as the spinal cord, medulla, and cerebral cortex and in the peripheral tissues (lymphocytes, fibroblasts) in ALS patients and normal control subjects. FMO expression was generally higher in tissues from ALS subjects than in control tissues, with the largest increases in FMO expression detected in the spinal cord. In peripheral tissues, the FMO mRNA level was found to be lower compared with FMO expression in brain tissue, and no differences were detected between ALS patients and the control tissue. FMO and PON gene expression was low in peripheral tissues. In contrast to FMO5 expression, the PON2 gene was down-regulated in ALS patients compared to the controls. Because FMO and PON are involved in the detoxication processes and their functional activity to bioactivate chemicals to toxins has been documented, the data herein suggest that environmental toxin exposure may play a role in a subset of individuals who contract ALS by altering FMO and PON gene expression. Although the precise pathogenic link is presently unknown, these findings suggest a role at FMO and PON genes in the development of ALS.

Effects of salinity acclimation on the pesticide-metabolizing enzyme flavin-containing monooxygenase (FMO) in rainbow trout (Oncorhynchus mykiss)

Thioether-containing pesticides are more toxic in certain anadromous and catadromous fish species that have undergone acclimation to hypersaline environments. Enhanced toxicity has been shown to be mediated through the bioactivation of these xenobiotics by one or more flavin-containing monooxygenases (FMOs), which are induced by hyperosmotic conditions. To better understand the number of FMO genes that may be regulated by hyperosmotic conditions, rainbow trout (Oncorhynchus mykiss) were maintained and acclimated to freshwater (<0.5g/L salinity) and to 18g/L salinity. The expression of 3 different FMO transcripts (A, B and C) and associated enzymatic activities methyl p-tolyl sulfoxidation (MTSO) and benzydamine N-oxigenation (BZNO) were measured in four tissues. In freshwater-acclimated organisms FMO catalytic activities were as follows: liver>kidney>gills=olfactory tissues; in hypersaline-acclimated animals activities were higher in liver>gills>olfactory tissues>kidney. Acclimation to 18g/L caused a significant induction in the stereoselective formation of R-MTSO in gill. In olfactory tissues, stereoselective (100%) formation of S-MTSO was observed and was unaltered by acclimation to hypersaline water. When specific transcripts were evaluated, salinity-acclimation increased FMO A in liver (up to 2-fold) and kidney (up to 3-fold) but not in olfactory tissues and gills. FMO B mRNA was significantly down-regulated in all tissues, and FMO C was unchanged by hypersaline acclimation. FMO B and C failed to correlate with any FMO catalytic activity, but FMO A mRNA expression linearly correlated to both FMO catalytic activities (MTSO and BZNO) in liver (r2=0.92 and r2=0.88) and kidney microsomes (r2=0.93 and r2=90). FMO A only correlated with MTSO activity in gills (r2=0.93). These results indicate unique tissue specific expression of FMO genes in salmonids and are consistent with salinity-mediated enhancement of thioether-containing pesticide bioactivation by FMO which may occur in liver or kidney after salinity acclimation.

Enhanced indirubin production in recombinant Escherichia coli harboring a flavin-containing monooxygenase gene by cysteine supplementation

In our previous study, a batch fermentation of recombinant Escherichia coli DH5α cells harboring the fmo gene from Methylophaga aminisulfidivorans MPT produced indirubin (5.0mg/L) and indigo (920mg/L) in a 5L fermenter containing tryptophan medium (2g/L tryptophan, 5g/L yeast extract, 10g/L NaCl). In this study, it was found that indirubin production greatly increased when 0.36g/L cysteine was added to the tryptophan medium, although cysteine inhibited the growth of the recombinant E. coli harboring the fmo gene. However, the addition of cysteine did not inhibit the expression level and activity of FMO in the cell. Indigo was synthesized by the dimerization of two 3-hydroxyindole molecules under the non-enzymatic reaction. Cysteine influenced the regioselectivity of FMO and enhanced the synthesis of 2-hydroxyindole instead of 3-hydroxyindole, which might function to increase indirubin production. The optimal culture conditions for indirubin production in tryptophan medium were determined from the response surface methodology analysis: 2g/L tryptophan, 5g/L yeast extract, 10g/L NaCl, 0.36g/L (3mM) cysteine, pH 8.0 at 35°C. Under these conditions, the recombinant E. coli cells were capable of producing 223.6mg/L of indirubin from 2g/L of tryptophan. The intracellular accumulation of the indirubin crystals might stress the cell, which may be a main reason for the poor growth of the recombinant E. coli pBlue 1.7.

Genome-wide association identifies genetic variants associated with lentiform nucleus volume in N = 1345 young and elderly subjects

Deficits in lentiform nucleus volume and morphometry are implicated in a number of genetically influenced disorders, including Parkinson's disease, schizophrenia, and ADHD. Here we performed genome-wide searches to discover common genetic variants associated with differences in lentiform nucleus volume in human populations. We assessed structural MRI scans of the brain in two large genotyped samples: the Alzheimer's Disease Neuroimaging Initiative (ADNI; N = 706) and the Queensland Twin Imaging Study (QTIM; N = 639). Statistics of association from each cohort were combined meta-analytically using a fixed-effects model to boost power and to reduce the prevalence of false positive findings. We identified a number of associations in and around the flavin-containing monooxygenase (FMO) gene cluster. The most highly associated SNP, rs1795240, was located in the FMO3 gene; after meta-analysis, it showed genome-wide significant evidence of association with lentiform nucleus volume (P MA = 4.79 × 10(-8)). This commonly-carried genetic variant accounted for 2.68% and 0.84% of the trait variability in the ADNI and QTIM samples, respectively, even though the QTIM sample was on average 50years younger. Pathway enrichment analysis revealed significant contributions of this gene to the cytochrome P450 pathway, which is involved in metabolizing numerous therapeutic drugs for pain, seizures, mania, depression, anxiety, and psychosis. The genetic variants we identified provide replicated, genome-wide significant evidence for the FMO gene cluster's involvement in lentiform nucleus volume differences in human populations.

Association of rs10912745 and rs4916375 polymorphisms located in the cluster of flavin-containing monooxygenase genes, with ischaemic cardioembolic stroke

Twenty-one SNPs located in the cluster of genes FMO1-FMO2-FMO3-FMO4 and adjacent areas were analyzed in the patients with ischaemic cardioembolic stroke and in the control group. Significant differences between these samples were found in the distribution of genotype and allele differences in two polymorphic loci, rs10912745 and rs4916375. It was shown that these polymorphic loci are associated with the risk of ischaemic cardioembolic stroke development.

Genotyping of flavin-containing mono-oxygenase 3 (FMO3) gene by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and mismatch amplification mutation assay (MAMA-PCR) in chickens

2Institute of Animal Science and Veterinary Medicine, Shanxi Academy of Agricultural Science, 030000, Taiyuan, China. Accepted 13 December, 2011 Fishy smelling off-flavour of chicken meat is caused by a mutation ( FMO3 c.985A>T ) in exon 7 of flavincontaining mono-oxygenase 3 (FMO3) gene. Comparative analysis of the causative mutation in FMO3 gene in different chickens using two different methods was reported. Polymerase chain reactionrestriction fragment length polymorphism (PCR-RFLP) method was based on general primers, whereas another employed method was the mismatch amplification mutation assay (MAMA) primers. Seventy (22.88%) of the 306 samples were genotyped using PCR-RFLP, 194 (63.40%) of the 306 samples were genotyped using MAMA-PCR and 42 (13.72%) of the 306 samples were genotyped by both of PCR-RFLP and MAMA-PCR and genotyping data were validated by DNA sequencing. The results show that the genotyping accuracy for PCR-RFLP was 90.48% in AA, 85.42% in AT and 95.35% in TT and 100% in AA, 98% in AT and 96% in TT for MAMA-PCR. This study suggests that the MAMA-PCR is more effective, accurate, easy, fast, efficient and reliable than PCR-RFLP.

Flavin-Containing Monooxygenase mRNA Levels are Up-Regulated in ALS Brain Areas in SOD1-Mutant Mice

Flavin-containing monooxygenases (FMOs) are a family of microsomal enzymes involved in the oxygenation of a variety of nucleophilic heteroatom-containing xenobiotics. Recent results have pointed to a relation between Amyotrophic Lateral Sclerosis (ALS) and FMO genes. ALS is an adult-onset, progressive, and fatal neurodegenerative disease. We have compared FMO mRNA expression in the control mouse strain C57BL/6J and in a SOD1-mutated (G93A) ALS mouse model. Fmo expression was examined in total brain, and in subregions including cerebellum, cerebral hemisphere, brainstem, and spinal cord of control and SOD1-mutated mice. We have also considered expression in male and female mice because FMO regulation is gender-related. Real-Time TaqMan PCR was used for FMO expression analysis. Normalization was done using hypoxanthine-guanine phosphoribosyl transferase (Hprt) as a control housekeeping gene. Fmo genes, except Fmo3, were detectably expressed in the central nervous system of both control and ALS model mice. FMO expression was generally greater in the ALS mouse model than in control mice, with the highest increase in Fmo1 expression in spinal cord and brainstem. In addition, we showed greater Fmo expression in males than in female mice in the ALS model. The expression of Fmo1 mRNA correlated with Sod1 mRNA expression in pathologic brain areas. We hypothesize that alteration of FMO gene expression is a consequence of the pathological environment linked to oxidative stress related to mutated SOD1.

Bio-indigo production by two types of fermentation systems using recombinant E. coli cells harboring a flavin-containing monooxygenase gene ( fmo)

Bio-indigo production by two types of fermentation systems using recombinant E. coli cells harboring a flavin-containing monooxygenase gene ( fmo)

Interactions of flavin containing monooxygenase 3 (FMO3) genotype and feeding of choline and rapeseed cake on the trimethylamine (TMA) content in egg yolks of laying hens

To evaluate the relationship between the FMO3 genotype and dietary trimethylamine (TMA) precursors on TMA-N content in eggs, laying hens were fed with choline (500 and 4000 mg/kg diet) and, in a later trial, with rapeseed cake (0, 100 and 300 g/kg diet). Hens were genotyped for non-synonymous A/T polymorphism at position nt1034 of the cDNA of the FMO3 gene, which is associated with elevated TMA-levels in egg yolks. Genotypes were equally distributed among the experimental diets. TMA was extracted from yolks and measured as TMA-N by a colorimetric method. As expected, hens of the TT-genotype had high amounts of TMA-N in egg yolks when fed either 4000 mg choline/kg diet or the rapeseed cake diet, while hens of the AA-genotype showed low levels of TMA-N. Heterozygous AT-hens had a slightly increased level of TMA-N when fed with 4000 mg choline/kg diet but not with the rapeseed cake feed. Rapeseed cake can be fed to AA and AT hens without leading to fishy tainted eggs.

Novel variants of the human flavin-containing monooxygenase 3 ( FMO3) gene associated with trimethylaminuria

The disorder trimethylaminuria (TMAu) often manifests itself in a body odor for individuals affected. TMAu is due to decreased metabolism of dietary-derived trimethylamine (TMA). In a healthy individual, 95% or more of TMA is converted by the flavin-containing monooxygenase 3 (FMO3, EC 1.14.13.8) to non-odorous trimethylamine N-oxide (TMA N-oxide). Several single nucleotide polymorphisms (SNPs) of the FMO3 gene have been described and result in an enzyme with decreased or abolished functional activity for TMA N-oxygenation thus leading to TMAu. Herein, we report two novel mutations observed from phenotyping and genotyping two self-reporting individuals. Sequence analysis of the exon regions of the FMO3 gene of a young woman with severe TMAu revealed heterozygous mutations at positions 187 (V187A), 158 (E158K), 308 (E308G), and 305 (E305X). Familial genetic analysis showed that the E158K/V187A/E308G derived from the same allele from the mother, and the E305X was derived from the father. FMO3 variants V187A and V187A/E158K were characterized for oxygenation of several common FMO3 substrates (i.e., 5- and 8-DPT, mercaptoimidazole (MMI), TMA, and sulindac sulfide) and for its thermal stability. Our findings show that with the combination of V187A/E158K mutations in FMO3, the enzyme activity is severely affected and possibly contributes to the TMAu observed. In another study, genotyping analysis of a 17 year old female revealed a mutation that caused a frame shift after K415 and resulted in a protein variant with only 486 amino acid residues that was associated with severe TMAu.

Individual Differences of Drug-metabolizing Enzymes as Determinants for the Metabolic Fate of Chemicals-A Study of Trimethylamine and Flavin-containing Monooxygenase 3-

Individual differences of drug-metabolizing enzymes are important determinants for the metabolic fate of chemicals. This article focuses on polymorphic human flavin-containing monooxygenase 3 (FMO3) and dietary-derived trimethylamine. Malodorous trimethylamine is generally converted to odorless trimethylamine N-oxide by liver microsomal FMO3. Trimethylaminuria is caused by functional disorder of FMO3. In this study mutations of the FMO3 gene were examined in self-reported Japanese trimethylaminuria subjects that showed low FMO3 metabolic capacity in urine tests. Nine novel polymorphisms in the FMO3 gene were discovered in self-reported Japanese volunteers. Functional analyses of recombinant FMO3 proteins suggested that these FMO3 gene mutations were one of the causal factors for decreased FMO3 function resulting in trimethylaminuria. Inter-individual variations of FMO3-mediated microsomal oxygenation activities, levels of FMO3 protein and FMO3 mRNA, and its modification in liver microsomes from Japanese samples were observed. Both genetic polymorphisms in the 5'-upstream of the FMO3 gene and some hormonal changes related to menstruation may be causal factors for inter- and/or intra- individual expression levels of FMO3. To assess the palliative cares, it was found that absorbed levels of trimethylamine in vivo would be possibly controlled by selection of precursor foods like fish containing a variety of trimethylamine amounts. These lines of evidence suggest that individual differences of FMO3 are important determinants for the metabolic fate of dietary-derived trimethylamine.