Mitochondrial proton leak in brown adipose tissue mitochondria of Ucp1-deficient mice is GDP insensitive

We have analyzed characteristics of mitochondrial proton leak from brown ad ipose tissue (BAT) of Ucp1-deficient mice and normal controls and conducted the first top-down metabolic control analysis of oxidative phosphorylation in BAT mitochondria. Because purine nucleotides inhibit UCP1 and because U CP2 and the long form of UCP3 have putative purine nucleotide-binding regio ns, we predicted that proton leak in BAT mitochondria from Ucp1-deficient m ice would be sensitive to GDP. On the contrary, although control over mitoc hondrial oxygen consumption and proton leak reactions at state 4 are strong ly affected by 1 mM GDP in mitochondria from normal mice, there is no effec t in UCP1-deficient mitochondria. In the presence of GDP, the overall kinet ics of proton leak were not significantly different between Ucp1-deficient mice and controls. In its absence, state 4 respiration in normal BAT mitoch ondria was double that in its presence. Leak-dependent oxygen consumption w as higher over a range of membrane potentials in its absence than in its pr esence. Thus proton leak, potentially including that through UCP2 and UCP3, is GDP insensitive. However, our measurements were made in the presence of albumin and may not allow for the detection of any fatty acid-induced UCP- mediated leak; this possibility requires investigation.