Respiratory chain energy conservation in the methylotrophic bacterium Methylophilus methylotrophus
M.methylotrophus is an obligately aerobic, Gram negative methylo- trophic bacterium which grows preferentially on methanol as the carbon and energy source, and uses the ribulose monophosphate pathway for carbon assimilation. This organism is used for single-cell protein production in the I.C.I. 'PRUTEEN' process. The composition and sequential organisation of the respiratory chain of M.methylotrophus have been studied, and both kinetic (→ H+/O, → K+/O quotients) and thermodynamic (Δμ⁻ H+, ΔGp) parameters of energy conservation have been determined. In addition, the effect of the growth conditions on some of these parameters has been investigated. The respiratory chain of M.methylotrophus was found to branch at the level of cytochrome c to two terminal oxidases, cytochromes aa3 and o. Methanol is oxidized via a methanol dehydrogenase which donates reducing equivalents to the respiratory chain at the level of cytochrome c, as in other methylotrophs. Proton and charge translocation stoicheiometries indicate the presence of three energy conserving sites between NADH and oxygen, each of which translocates two charges; only the third coupling site, which appears to function by a redox arm mechanism, is involved in respiration from methanol. M.methylotrophus was found to sustain a ?Gp, during respiration from methanol, of approximately -45 kJ/mol, but the Δμ⁻ H+ varied with the reaction conditions such that apparent values of the ? H+/ATP quotient ranging from 2.6 to 4.1 g-ion H+/mol ATP were obtained. It was concluded that the proton current, in this organism, is at least partially localised, and theoretical growth calculations suggest that the true value of the ? H+/ATP quotient is probably 2 g-ion H+/mol ATP. On this basis, the ATP/O quotients for respiration from NADH and methanol are likely to be 3 and 1 mol ATP/g-atom O, respectively. There was no evidence that the low growth yields of methanol-excess cultures could be explained by a reduced efficiency of respiratory chain energy conservation.