The 1.3 Mbp genome of HTCC2181, a member of the abundant OM43 clade of coastal bacterioplankton, suggested it is an obligate methylotroph. Preliminary experiments demonstrated that methanol and formaldehyde, but not other common C1 compounds such as methylamine, could support growth. Methanol concentrations in seawater are reportedly < 100 nM, suggesting either that the flux of methanol through plankton pools is very rapid, or that methanol may not be the primary growth substrate for HTCC2181. Therefore, we investigated the apparent extreme substrate range restriction of HTCC2181 in greater detail. Growth rate and maximum cell density of HTCC2181 increased with methanol concentration, yielding aKs value of 19 μM. In contrast, no growth was observed in the presence of the methylated (C1) compounds, methyl chloride, trimethylamine-oxide (TMAO) or dimethylsulfoniopropionate (DMSP) when they were the sole substrates. However, growth rate, maximum cell density and cellular ATP content were significantly enhanced when any of these methylated compounds were provided in the presence of a limiting concentration of methanol. These observations fit a model in which the metabolic intermediate formaldehyde is required for net carbon assimilation, allowing C1 substrates that do not produce a formaldehyde intermediate to be oxidized for energy, but not assimilated into biomass. Rates of methanol and TMAO oxidation and assimilation were measured with 14C-radiolabelled compounds in cultures of HTCC2181 and seawater microbial communities collected off the Oregon coast. The results indicated that in nature as well as in culture, C1 substrates are partitioned between those that are mainly oxidized to produce energy and those that are assimilated. These findings indicate that the combined fluxes of C1 compounds in coastal systems are sufficient to support significant populations of obligate methyltrophs by a metabolic strategy that involves the synergistic metabolism of multiple C1 compounds.