Eduardo Rial

The regulation of the proton conductance of brown fat mitochondria. Identification of functional and non-functional nucleotide-binding sites.

The binding of purine nucleotides to intact brown fat mitochondria is re‐examined. In addition to the previously reported high affinity binding site, a low‐affinity site is found, which requires several minutes to saturate. Only the high affinity site is functional in regulating the proton and halide permeabilities of the mitochondria. The low affinity site can introduce errors in the use of nucleotide binding to quantitate the M r 32 000 uncoupling protein unique to these mitochondria.

Chemical modification of the brown fat mitochondrial uncoupling protein with tetranitromethane

Tetranitromethane reacts with the uncoupling protein of intact brown fat mitochondria. The chloride permeability in the absence of the inhibitory nucleotide GDP is not affected, but the affinity with which GDP binds is decreased, and the coupling between binding of nucleotide and inhibition of chloride permeation is broken.

On the Mechanism of Transport by the Uncoupling Protein from Brown Adipose Tissue Mitochondria

The uncoupling protein enables brown adipose tissue mitochondria to uncouple respiration from ATP synthesis under thermogenic conditions by catalyzing re-entry of protons extruded by the respiratory chain. Two ligands, purine nucleotides and fatty acids, interact specifically with the protein to modulate its transport activity (for review see Rial & Nicholls, 1987).

The Function of the Uncoupling Protein in the Intact Brown Fat Cell

When stimulated by the sympathetic nervous system, the brown fat of a cold-adapted, maximally thermogenic, animal is capable of respiring at 60 times the rate of an equivalent mass of liver (Foster and Frydman, 1978). The molecular basis for this spectacular rate of respiration was established some years ago, when we described the presence in the brown fat mitochondrial inner membrane of a unique 32kDa protein (Heaton et al., 1978). This “uncoupling protein” allows the protons extruded by the respiratory chain to re-enter the matrix bypassing the ATP synthase and the kinetic restraints of respiratory control. This paper will address two questions: firstly, what is the evidence that the uncoupling protein is functional in the intact cell? Secondly what is the nature of the cytoplasmic second messenger which regulates the activity of this energy dissipating pathway?