Ruegeria pomeroyi DSS-3 possesses two general pathways for metabolism of dimethylsulphoniopropionate (DMSP), an osmolyte of algae and abundant carbon source for marine bacteria. In the DMSP cleavage pathway, acrylate is transformed into acryloyl-CoA by propionate-CoA ligase (SPO2934) and other unidentified acyl-CoA ligases. Acryloyl-CoA is then reduced to propionyl-CoA by AcuI or SPO1914. Acryloyl-CoA is also rapidly hydrated to 3-hydroxypropionyl-CoA by acryloyl-CoA hydratase (SPO0147). A SPO1914 mutant was unable to grow on acrylate as the sole carbon source, supporting its role in this pathway. Similarly, growth on methylmercaptopropionate, the first intermediate of the DMSP demethylation pathway, was severely inhibited by a mutation in the gene encoding crotonyl-CoA carboxylase/reductase, demonstrating that acetate produced by this pathway was metabolized by the ethylmalonyl-CoA pathway. Amino acids and nucleosides from cells grown on 13C-enriched DMSP possessed labelling patterns that were consistent with carbon from DMSP being metabolized by both the ethylmalonyl-CoA and acrylate pathways as well as a role for pyruvate dehydrogenase. This latter conclusion was supported by the phenotype of a pdh mutant, which grew poorly on electron-rich substrates. Additionally, label from [13C-methyl] DMSP only appeared in carbons derived from methyl-tetrahydrofolate, and there was no evidence for a serine cycle of C-1 assimilation.