The stoicheiometries of respiration-linked proton translocation in Methylophilus methylotrophus were determined by using both the oxygen-pulse and initial-rate methods. The latter has also been used to determine leads to charge/O quotients (measured as yield K+/O quotients) in order to ascertain whether the leads to H+/O quotients might be underestimated by H+/anion symport. The results suggest that 6H+/O are translocated during NADH oxidation, and that 2H+/O are translocated during the oxidation of methanol to formaldehyde. There was no evidence for underestimation of the leads to H+/O quotients due to H+/anion symport, except by the movement of formic acid during formate oxidation. By comparing these results with the known growth efficiencies of this organism, an leads to H+/ATP quotient of close to 2 g-ions of H+/mol of ATP can be calculated. It is proposed that the respiratory chain in Methylophilus methylotrophus is arranged such that there are three sites of energy conservation for NADH oxidation, each translocating 2H+ and each linked to the synthesis of one molecule of ATP. Only the third site of energy conservation is involved in methanol oxidation.

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