Partitioning of 14C derived from [14C]-methane into biomass and CO2 over 1 h is shown in Fig. 2. Under control conditions PI3 kinase pathway (i.e. in the absence of Hg2+), 61 ± 4% of 14C is assimilated and 23 ± 3% is oxidized to CO2 per hour, with the remainder presumably not oxidized or in solution either as methane or as soluble metabolites. Foster & Davis (1966) found the partitioning of methane by M. capsulatus TexasT to be 16% to CO2, 63% to biomass and 21% to ‘soluble carbon’. Leak and Dalton (1986a, b) comment that growth yields in M. capsulatus (Bath) are variable with growth conditions, but values between 19% and 70% of methane–carbon
assimilated are reported, with the remaining 71% and 30% of methane–carbon going to CO2 and soluble intermediates. In the presence of 10 mM HgCl2, almost all methane (39.6 ± 0.9 nmol) was converted to CO2 within 30 min with no assimilation and apparently minimal leakage of soluble metabolites (determined by difference). After 1 h incubation, the medium in HgCl2-containing flasks had taken
on a greyish tone, which was also evident in harvested cells. This was presumed to be because of elemental mercury adsorbing onto particulates – total reduction of the 500 μmol Hg2+ present would release approximately 8 μL elemental mercury per flask. No greying of the medium was found in killed controls. Given the rapid nature of the oxidation of methane to CO2 in the presence of Hg2+ with
no lag phase in which carbon was assimilated, see more it is assumed that the regulation of this process occurs immediately, at the protein level. The oxidation of methane to CO2 in M. capsulatus (Bath) proceeds via methanol, formaldehyde almost and formate. Most of the formaldehyde and, to some extent, formate are assimilated to biomass via the Quayle (ribulose monophosphate, RuMP) pathway with some formate oxidized to CO2 to generate reducing equivalents to meet the energy demand of the cell. Mercuric reductase activity would require NAD(P)H and this demand could be met in cells by oxidizing all available methane to CO2, generating NADH from the terminal oxidation of formate by formate dehydrogenase (EC 1.2.1.2). For the cytochrome c oxidase pathway, reduced cytochrome c is required as the cofactor for the oxidase (EC 1.9.3.1), which must be produced in vivo at the expense of reducing equivalents, which could be obtained by the total oxidation of methane to CO2. Given that the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO, green form, EC 4.1.1.39) activity in M. capsulatus (Bath) when grown on methane (Taylor et al., 1981; Stanley & Dalton, 1982), some of the CO2 produced could be reassimilated, but this is not the case when Hg2+ and Hg are present, which would indicate that one of these species inhibits RuBisCO activity, as is the case in Nitrosomonas sp. K1 (Hatayama et al., 2000).