title: Phenotyping of flavin-containing monooxygenase using caffeine metabolism and genotyping of FMO3 gene in a Korean population.
abstract: Flavin-containing monooxygenase (FMO) activity was determined in 82 Korean volunteers by taking molar concentration ratio of theobromine and caffeine present in the 1 h urine (between 4 and 5 h) samples collected after administration of a cup of coffee containing 110 mg of caffeine. Among 82 volunteers, there were 19 women and 63 men (30 smokers and 52 non-smokers). Volunteers were divided into two groups comprising low (0.53-2.99) and high (3.18-11.95) FMO activities separated by an antimode of 3.18. Peripheral bloods were sampled from these volunteers and their genomic DNAs were amplified by polymerase chain reaction with oligonucleotides designed from intronic sequences of human FMO3 gene. Comparing nucleotide sequences of the amplified FMO3 gene originating from randomly selected individuals with low and high FMO activities, nine point mutations were identified in the open reading frame sequences. Among these nine mutations, three FMO3 mutant types (FMO3/Stop148, Lys158 and Gly308) were selected and correlated with FMO activities observed in our Korean population. A rare FMO3/Stop148 mutant allele originating from FMO3/Gly148 occurred by substitution of G442T in exon 4 and yielded a premature TGA stop codon. The stop codon was detected in one individual having the second lowest FMO activity and he had the mutation in heterozygous state. In a pedigree study, he was found to have inherited the mutation from his mother who also had a heterozygous stop codon and equally low FMO activity. In our volunteers, two other common mutations were detected in exons 4 and 7. The one in exon 4 resulted from a G472A change eliminating a HinfI restriction site and produced an amino acid substitution from Glu158 to Lys. The other mutation in exon 7 resulted from an A923G change generating a DraII restriction site and produced a non-conservative replacement of Glu308 to Gly. Based on the secondary structure maps of FMO3 enzyme proteins for these two mutant types, FMO3/Gly308 mutation transformed the helix structure into a sheet shape and indicated that dysfunctional FMO3 may be produced. FMO3/Lys158 mutation did not alter the secondary structure. Approximately 80% of volunteers with homozygous and/or heterozygous mutations on either one or two of these mutations had low FMO activities. Thus, individuals with these FMO3 gene mutations may have defective metabolic activity for many clinically used drugs and dietary plant alkaloids which are oxidized primarily by hepatic FMO3.
authors.affiliation: Department of Pharmacology and Toxicology, College of Medicine, Inha University, Inchon, Korea.
authors.forename: C S
authors.initials: CS
authors.lastname: Park
authors.forename: W G
authors.initials: WG
authors.lastname: Chung
authors.forename: J H
authors.initials: JH
authors.lastname: Kang
authors.forename: H K
authors.initials: HK
authors.lastname: Roh
authors.forename: K H
authors.initials: KH
authors.lastname: Lee
authors.forename: Y N
authors.initials: YN
authors.lastname: Cha
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002110:Caffeine / Q000378:metabolism* / Q000652:urine; D018388:Codon, Terminator; D017931:DNA Primers; D005260:Female; D005838:Genotype; D006801:Humans; D007723:Korea; D008297:Male; D009154:Mutation; D010105:Oxygenases / Q000235:genetics* / Q000378:metabolism; D010375:Pedigree; D010641:Phenotype; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D013805:Theobromine / Q000378:metabolism / Q000652:urine
chemical_list: D018388:Codon, Terminator; D017931:DNA Primers; D002110:Caffeine; D010105:Oxygenases; C020066:dimethylaniline monooxygenase (N-oxide forming); D013805:Theobromine