title: Identification and characterization of trimethylamine N-oxide (TMAO) demethylase and TMAO permease in Methylocella silvestris BL2.
abstract: Methylocella silvestris, an alphaproteobacterium isolated from a forest soil, can grow on trimethylamine N-oxide (TMAO) as a sole nitrogen source; however, the molecular and biochemical mechanisms underpinning its growth remain unknown. Marker-exchange mutagenesis enabled the identification of several genes involved in TMAO metabolism, including Msil_3606, a permease of the amino acids-polyamine (APC) superfamily, and Msil_3603, consisting of an N-terminal domain of unknown function (DUF1989) and a C-terminal tetrahydrofolate-binding domain. Null mutants of Msil_3603 and Msil_3606 can no longer grow on TMAO. Purified Msil_3603 from recombinant Escherichia coli can convert TMAO to dimethylamine and formaldehyde (1 TMAO → 1 dimethylamine + 1 formaldehyde), confirming that it encodes a bona fide TMAO demethylase (Tdm). Tdm of M. silvestris and eukaryotic Tdms have no sequence homology and contrasting characteristics. Recombinant Tdm of M. silvestris appears to be hexameric, has a high affinity for TMAO (Km = 3.3 mM; Vmax = 21.7 nmol min(-1)  mg(-1) ) and only catalyses demethylation of TMAO and a structural homologue, dimethyldodecylamine N-oxide. Our study has contributed to the understanding of the genetic and biochemical mechanisms for TMAO degradation in M. silvestris.
authors.affiliation: School of Life Sciences, University of Warwick, Coventry, CV4 7AL, UK.
authors.forename: Yijun
authors.initials: Y
authors.lastname: Zhu
authors.forename: Eleanor
authors.initials: E
authors.lastname: Jameson
authors.forename: Rosemary A
authors.initials: RA
authors.lastname: Parslow
authors.forename: Ian
authors.initials: I
authors.lastname: Lidbury
authors.forename: Tiantian
authors.initials: T
authors.lastname: Fu
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Dafforn
authors.forename: Hendrik
authors.initials: H
authors.lastname: Schäfer
authors.forename: Yin
authors.initials: Y
authors.lastname: Chen
mesh_terms: D000446:Aldehyde-Lyases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D020561:Alphaproteobacteria / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D008744:Methylamines / Q000378:metabolism*; D016296:Mutagenesis
chemical_list: D026901:Membrane Transport Proteins; D008744:Methylamines; D000446:Aldehyde-Lyases; C023188:trimethylamine-oxide aldolase; C005855:trimethyloxamine