Control of the effective P/O ratio of oxidative phosphorylation in liver mitochondria and hepatocytes

Abstract

The control exerted by substrate oxidation reactions, by ATP turnover and by the proton leak over the oxygen consumption rate, the phosphorylation rate, the proton leak rate and the protonmotive force (delta p) in isolated rat liver mitochondria under a range of conditions between non-phosphorylating (State 4) and maximum phosphorylation (State 3) was investigated by using the top-down approach of metabolic control analysis. The experiments were carried out with saturating concentrations of the substrates succinate, glutamate with malate, or pyruvate with malate. The distribution of control was very similar with each of the three substrates. The effective P/O ratio (i.e. not corrected for leak reactions) was also measured; it varied from zero in State 4 to 80-90% of the maximum theoretical P/O ratio in State 3. Under most conditions control over the effective P/O ratio was shared between proton leak (which had negative control) and the phosphorylating subsystem (which had roughly equal positive control); near State 4, substrate oxidation reactions also acquired some control over this ratio. In resting hepatocytes the effective P/O ratio was only 50% of its maximum theoretical value, corresponding to an effective P/O ratio of only 1.3 for complete oxidation of glucose. The effective P/O ratio for intracellular mitochondrial oxygen consumption was 64% of the maximum value. The control coefficient of the mitochondrial proton leak over the effective P/O ratio in cells was -0.34; the control coefficient of phosphorylation reactions over this ratio was 0.31 and the control coefficient of substrate oxidation reactions over the ratio was 0.03, showing how the coupling efficiency in cells can respond sensitively to agents that change the proton leak or the ATP demand, but not to those that change substrate oxidation.

The nature and significance of nutritional adaptation

“…The greater part of the body's energy expenditure is on the synthesis of ATP rather than its utilization. About 25% of the energy involved in ATP synthesis is dissipated in the mitochondrial proton leak (Rolfe & Brown, 1997), the extent of which is reduced in hypothyroidism, so that it would be an advantage to be mildly hypothyroid (Brand et al, 1993). The only measurement of the PaO ratio in the intact human being that I know of is the ingenious and elegant study of Flatt et al (1984).…”

Fatty acid metabolism, energy expenditure and insulin resistance in muscle

“…In reality, coupling efficiency can vary significantly depending on changes in proton leak or ATP demand, but in cell systems at least, changes in substrate oxidation do not appear to influence the relationship between oxygen consumption and ATP synthesis (Brand et al 1993). …”

Mitochondrial uncoupling protein 2 in pancreatic β‐cells

“…This supports the idea that UCP2 normally contributes significantly to the proton conductance of the membrane and attenuates the rise in protonmotive force, ATP/ADP ratio and insulin secretion as glucose concentration increases. It is formally possible that UCP2 knockdown confers increased coupling efficiency not through decreased proton leak, but through increases in ATP turnover and/or glucose oxidation [33]. However, a decrease in levels of UCP2 lowers specific oxygen consumption rates in both trypsinized [31] and attached INS-1E cells (C. Affourtit, M. Jastroch and M. D. Brand, unpublished data) and in pancreatic islets [34].…”