Choline degradation in Paracoccus denitrificans: identification of sources of formaldehyde

Paracoccus denitrificans is a facultative methylotroph that can grow on methanol and methylamine as sole sources of carbon and energy. Both are oxidized to formaldehyde and then to formate, so growth on C1 substrates induces the expression of genes encoding enzymes required for the oxidation of formaldehyde and formate. This induction involves a histidine kinase response regulator pair (FlhSR) that is likely triggered by formaldehyde. Catabolism of some complex organic substrates (e.g., choline and L-proline betaine) also generates formaldehyde. Thus, flhS and flhR mutants that fail to induce expression of the formaldehyde catabolic enzymes cannot grow on methanol, methylamine, and choline. Choline is oxidized to glycine via glycine betaine, dimethylglycine, and sarcosine. By exploring flhSR growth phenotypes and the activities of a promoter and enzyme known to be upregulated by formaldehyde, we identify the oxidative demethylations of glycine betaine, dimethylglycine, and sarcosine as sources of formaldehyde. Growth on glycine betaine, dimethylglycine, and sarcosine is accompanied by the production of up to three, two, and one equivalents of formaldehyde, respectively. Genetic evidence implicates two orthologous monooxygenases in the oxidation of glycine betaine. Interestingly, one of these appears to be a bifunctional enzyme that also oxidizes L-proline betaine (stachydrine). We present preliminary evidence to suggest that growth on L-proline betaine induces expression of a formaldehyde dehydrogenase distinct from the enzyme induced during growth on other formaldehyde-generating substrates.IMPORTANCEThe bacterial degradation of one-carbon compounds (methanol and methylamine) and some complex multi-carbon compounds (e.g., choline) generates formaldehyde. Formaldehyde is toxic and must be removed, which can be done by oxidation to formate and then to carbon dioxide. These oxidations provide a source of energy; in some species, the CO2 thus generated can be assimilated into biomass. Using the Gram-negative bacterium Paracoccus denitrificans as the experimental model, we infer that oxidation of choline to glycine generates up to three equivalents of formaldehyde, and we identify the three steps in the catabolic pathway that are responsible. Our work sheds further light on metabolic pathways that are likely important in a variety of environmental contexts.