Nat. Microbiol. 4, 155–163 (2019)

Elevated levels of circulating trimethylamine N-oxide (TMAO) are associated with multiple human diseases. TMAO arises from the oxidation of trimethylamine (TMA), which can be produced from choline by certain gut microorganisms harboring the choline utilization (cut) operon. To understand how these microbes obtain choline, Chittim et al. first tested representative gut bacteria for their ability to process choline-containing substrates, finding several strains that could cleave the membrane lipid phosphatidylcholine (PC) to choline. Bioinformatic analysis led to the identification of a phospholipase D (PLD)-encoding gene present only in PC-metabolizing strains. Deletion of this gene in an Escherichia coli isolate abolished the strain’s ability to produce choline or TMA from PC and to use PC as a carbon source. In vitro biochemical characterization of the E. coli PLD validated PC as the enzyme’s preferred substrate, while further bioinformatic analyses and mutagenesis implicated an active site glutamate residue as important for substrate selectivity. As these PLDs are widespread among choline-utilizing gut bacteria, distinct from those PLDs found in pathogens, and insensitive to human PLD inhibitors, their identification may help in the development of new selective therapeutics.