Inter-relationships between proton electrochemical gradient, adenine-nucleotide phosphorylation potential and respiration, during substrate-level and oxidative phosphorylation by mitochondria from brown adipose tissue of cold-adapted guinea-pigs.

Abstract

The inter-relationships between respiratory rates, proton electrochemical gradients (ΔH+) and extra-mitochondrial adenine nucleotide phosphorylation potentials (ΔGp(out) are examined during oxidative and substrate-level phosphorylation by mitochondria from the brown-adipose tissue of cold-adapted guinea-pigs. In the absence of net ATP synthesis, ΔGp(out) is proportional to ΔH+-when the latter is varied from 230 mV to 190 mV, and is consistent with a stoichiometry of proton translocation for ATP synthesis ( H+/ATP) of 2.6. When there is a net production of ATP, ΔGp(out) falls below the level predicted from ΔH+. When ATP is generated by substrate-level phosphorylation, ΔGp(out) can be much greater than that predicted from ΔH+. In the absence of substrate-level phosphorylation, respiration is controlled by ΔH+, regardless of whether energy dissipation is varied by addition of proton translocators or by addition of extra-mitochondrial ATP-hydrolysing systems. In contrast, the rate of respiration coupled to substrate-level phosphorylation appears to be controlled by the internal adenine nucleotide phosphorylation potential ΔGp(in). The rate of ATP synthesis by oxidative phosphorylation is dependent in ΔGp(out). In the absence of net ATP synthesis, both oxidative and substrate-level phosphorylation can maintain a ΔGp(out) in excess of 580 mV (56 kJ/mol), while in the presence of sufficient proton translocator to achieve uncontrolled respiration, a ΔGp(out) of 500 mV (48 kJ/mol) can be maintained by oxidative phosphorylation. Under conditions designed to approximate to those pertaining in the brown adipocyte during non-shivering thermogenesis, oxidative phosphorylation alone appears to be adequate to maintain cellular ATP levels. It was not possible to confirm reports of a specific role which could be assigned to substrate-level phosphorylation in the regulation of energy-dissipation by these mitochondria.