Molecular cloning, sequencing, and expression in Escherichia coli of mouse flavin-containing monooxygenase 3 (FMO3): Comparison with the human isoform

J. Greg Falls, Nathan J. Cherrington, Kieran M. Clements, Richard M. Philpot, Patricia E. Levi, Randy L. Rose, Ernest Hodgson

Research output: Contribution to journalArticle

33 Scopus citations

Abstract

The sequence of mouse flavin-containing monooxygenase 3 (FMO3) was obtained from several clones isolated from a mouse liver cDNA library. The nucleotide sequence of mouse FMO3 was 2020 bases in length containing 37 bases in the 5' flanking region, 1602 in the coding region, and 381 in the 3' flanking region. The derived protein sequence consisted of 634 amino acids including the putative flavin adenine dinucleotide and NADP+ pyrophosphate binding sites (characteristic of mammalian FMOs) starting at positions 9 and 191, respectively. The mouse FMO3 protein sequence was 79 and 82% identical to the human and rabbit FMO3 sequences, respectively. Mouse FMO3 was expressed in Escherichia coli and compared to E. coli expressed human FMO3. The FMO3 proteins migrated with the same mobility (~58 kDa) as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. The expressed FMO3 enzymes (mouse and human forms) were sensitive to heat and reacted in a similar manner toward metal ions and detergent. Catalytic activities of mouse and human FMO3 were high toward the substrate methimazole; however, in the presence of trimethylamine and thioacetamide, FMO-dependent methimazole oxidation by both enzymes was reduced by greater than 85%. Other substrates which inhibited methimazole oxidation were thiourea and thiobenzamide and to a lesser degree N,N-dimethylaniline. When probed with mouse FMO3 cDNA, FMO3 transcripts were detected in hepatic mRNA samples from female mice, but not in samples from males. FMO3 was detected in mRNA samples from male and female mouse lung, but FMO3 message was not detected in mouse kidney sample from either gender. Results of immunoblotting confirmed the tissue- and gender-dependent expression of mouse FMO3.

Original languageEnglish (US)
Pages (from-to)9-18
Number of pages10
JournalArchives of Biochemistry and Biophysics
Volume347
Issue number1
DOIs
StatePublished - Nov 1 1997
Externally publishedYes

Keywords

  • FMO3
  • FMO3 expression
  • FMO3 sequence
  • Flavin-containing monooxygenase 3

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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