title: Carnitine metabolism to trimethylamine by an unusual Rieske-type oxygenase from human microbiota.
abstract: Dietary intake of L-carnitine can promote cardiovascular diseases in humans through microbial production of trimethylamine (TMA) and its subsequent oxidation to trimethylamine N-oxide by hepatic flavin-containing monooxygenases. Although our microbiota are responsible for TMA formation from carnitine, the underpinning molecular and biochemical mechanisms remain unclear. In this study, using bioinformatics approaches, we first identified a two-component Rieske-type oxygenase/reductase (CntAB) and associated gene cluster proposed to be involved in carnitine metabolism in representative genomes of the human microbiota. CntA belongs to a group of previously uncharacterized Rieske-type proteins and has an unusual "bridging" glutamate but not the aspartate residue, which is believed to facilitate intersubunit electron transfer between the Rieske center and the catalytic mononuclear iron center. Using Acinetobacter baumannii as the model, we then demonstrate that cntAB is essential in carnitine degradation to TMA. Heterologous overexpression of cntAB enables Escherichia coli to produce TMA, confirming that these genes are sufficient in TMA formation. Site-directed mutagenesis experiments have confirmed that this unusual "bridging glutamate" residue in CntA is essential in catalysis and neither mutant (E205D, E205A) is able to produce TMA. Taken together, the data in our study reveal the molecular and biochemical mechanisms underpinning carnitine metabolism to TMA in human microbiota and assign the role of this novel group of Rieske-type proteins in microbial carnitine metabolism.
authors.affiliation: School of Life Sciences and Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors.forename: Yijun
authors.initials: Y
authors.lastname: Zhu
authors.forename: Eleanor
authors.initials: E
authors.lastname: Jameson
authors.forename: Marialuisa
authors.initials: M
authors.lastname: Crosatti
authors.forename: Hendrik
authors.initials: H
authors.lastname: Schäfer
authors.forename: Kumar
authors.initials: K
authors.lastname: Rajakumar
authors.forename: Timothy D H
authors.initials: TD
authors.lastname: Bugg
authors.forename: Yin
authors.initials: Y
authors.lastname: Chen
mesh_terms: D040981:Acinetobacter baumannii / Q000378:metabolism; D002331:Carnitine / Q000378:metabolism*; D002852:Chromatography, Ion Exchange; D019295:Computational Biology; D004926:Escherichia coli; D006801:Humans; D008744:Methylamines / Q000378:metabolism; D064307:Microbiota / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D010088:Oxidoreductases / Q000378:metabolism*; D010105:Oxygenases / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet
chemical_list: D008744:Methylamines; D010088:Oxidoreductases; D010105:Oxygenases; C023336:trimethylamine; D002331:Carnitine
keywords: comparative genomics; gut microbiota; methylated amine metabolism