Energy conservation during aerobic growth in Paracoccus denitrificans

Abstract

Paracoccus denitrificans was aerobically grown in chemostat culture with succinate or gluconate as carbon source. Due to the presence of two phosphorylation sites in the respiratory chain and the absence of branching, theoretical P/O ratios of 1.71 and 1.82 were calculated for cells growing respectively with succinate and gluconate as carbon source. Using these data, 95% confidence intervals for the P/O ratio were determined, via a mathematical model, at 0.91-1.15 and 1.00-1.37 for sulphate-limited cultures, with respectively succinate and gluconate as carbon source. These results and measurements of P/O ratios in membrane particles and of proton translocation in whole cells have led to the conclusion that site I phosphorylation is affected under sulphate-limited conditions. Under conditions of carbon source-limitation the endogenous leads to H+/O ratio is about 7-8. Average values of 3.40 and 4.78 were respectively found for sulphate-limited succinate- and gluconate grown cells. For starved cells, oxidizing succinate as exogenous substrate, the leads to H+/O ratios were determined at about 3-4, independent of the growth limiting factor. It is concluded that the number of protons ejected per pair of electrons per energy-conserving site (leads to H+/site ratio) is about 3-4, instead of 2 as postulated by the chemiosmotic hypothesis.

Theoretical calculations on the influence of the inorganic nitrogen source on parameters for aerobic growth of microorganisms

“…A number of possibilities to allow the determination of YA MAx and the P/O ratio from aerobic yield studies have been outlined by Stouthamer (1977). In heterotrophic cells the absence of phosphorylation site III has been demonstrated (Imai, Asano and Sato, 1967;Knobloch, Ishaque and Aleem, 1971 ;van Verseveld and Stouthamer, 1976;Meijer et al, 1977).…”

“…The calculations have been performed with the theoretical value for YATP (15.4) for growth with succinate and ammonia (Stouthamer, 1973) and for an experimental value (YmP = 7.5) determined for P. denitrificans ; van Verseveld, Meijer and Stouthamer, 1977). The YmP values for growth with nitrate and nitrogen with various ATP/N/ratios were calculated as described earlier for growth with glucose ( Table 1).…”

Proton translocation coupled to ubiquinol oxidation in Paracoccus denitrificans

“…However, in experiments of very similar design, using rotenone instead of piericidin A to inhibit electron flow in the site 1 region, Meijer et a&. (7) observed an t H + :O ratio of 3.0 associated with succinate oxidation. The cells in these separate investigations were grown under almost identical conditions in continuous culture, and it is, therefore.…”

“…Hence, Meijer ef ak. (7) assumed that the cells in their experiments, which were grown aerobically in a chemostat under conditions of succinate limitation, were deficient with respect to coupling site 3 and concluded that the t H + :site ratio was equal to the stoichiometry of proton translocation associated with succinate oxidation ( tH+:site = t H + : 8 ratio = 3-4) in rotenonetreated cells. It would appear that one possible emplanation for the discrepancy between the results reported both in this paper and previously (6) and those of Meijer et at.…”

“…It would appear that one possible emplanation for the discrepancy between the results reported both in this paper and previously (6) and those of Meijer et at. (7) lies in the proton translocating nature of the terminal cytochrorne oxidase present in the cells used in the different studies (22). Perhaps a less likely explanation for the difference in the observed t H + : 8 ratios associated with succinate oxidation in these similar independent investigations relates to the concentrations of succinate employed in the different measurements of proton translocation.…”