Uncoupling protein-1 is not leaky

Biochim Biophys Acta. Jun-Jul 2010;1797(6-7):773-84. doi: 10.1016/j.bbabio.2010.04.007. Epub 2010 Apr 14.

Abstract

The activity of uncoupling protein-1 (UCP1) is rate-limiting for nonshivering thermogenesis and diet-induced thermogenesis. Characteristically, this activity is inhibited by GDP experimentally and presumably mainly by cytosolic ATP within brown-fat cells. The issue as to whether UCP1 has a residual proton conductance even when fully saturated with GDP/ATP (as has recently been suggested) has not only scientific but also applied interest, since a residual proton conductance would make overexpressed UCP1 weight-reducing even without physiological/pharmacological activation. To examine this question, we have here established optimal conditions for studying the bioenergetics of wild-type and UCP1-/- brown-fat mitochondria, analysing UCP1-mediated differences in parallel preparations of brown-fat mitochondria from both genotypes. Comparing different substrates, we find that pyruvate (or palmitoyl-L-carnitine) shows the largest relative coupling by GDP. Comparing albumin concentrations, we find the range 0.1-0.6% optimal; higher concentrations are inhibitory. Comparing basic medium composition, we find 125 mM sucrose optimal; an ionic medium (50-100 mM KCl) functions for wild-type but is detrimental for UCP1-/- mitochondria. Using optimal conditions, we find no evidence for a residual proton conductance (not a higher post-GDP respiration, a lower membrane potential or an altered proton leak at highest common potential) with either pyruvate or glycerol-3-phosphate as substrates, nor by a 3-4-fold alteration of the amount of UCP1. We could demonstrate that certain experimental conditions, due to respiratoty inhibition, could lead to the suggestion that UCP1 possesses a residual proton conductance but find that under optimal conditions our experiments concur with implications from physiological observations that in the presence of inhibitory nucleotides, UCP1 is not leaky.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adipose Tissue, Brown / drug effects
  • Adipose Tissue, Brown / metabolism
  • Animals
  • Apoptosis Regulatory Proteins / drug effects
  • Apoptosis Regulatory Proteins / metabolism
  • Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / pharmacology
  • Cattle
  • Culture Media
  • Electrochemistry
  • Energy Metabolism / drug effects
  • Glycerophosphates / pharmacology
  • Guanosine Diphosphate / pharmacology
  • In Vitro Techniques
  • Ion Channels / antagonists & inhibitors
  • Ion Channels / deficiency
  • Ion Channels / genetics
  • Ion Channels / metabolism*
  • Male
  • Membrane Potential, Mitochondrial / drug effects
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Mitochondrial Proteins / antagonists & inhibitors
  • Mitochondrial Proteins / deficiency
  • Mitochondrial Proteins / genetics
  • Mitochondrial Proteins / metabolism*
  • Oxygen Consumption / drug effects
  • Protons
  • Pyruvic Acid / metabolism
  • RNA-Binding Proteins
  • Ribosomal Proteins / drug effects
  • Ribosomal Proteins / metabolism
  • Serum Albumin, Bovine
  • Succinic Acid / metabolism
  • Uncoupling Agents / pharmacology
  • Uncoupling Protein 1

Substances

  • Apoptosis Regulatory Proteins
  • Culture Media
  • DAP3 protein, human
  • Glycerophosphates
  • Ion Channels
  • Mitochondrial Proteins
  • Protons
  • RNA-Binding Proteins
  • Ribosomal Proteins
  • UCP1 protein, human
  • Ucp1 protein, mouse
  • Uncoupling Agents
  • Uncoupling Protein 1
  • Guanosine Diphosphate
  • Serum Albumin, Bovine
  • Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone
  • Pyruvic Acid
  • alpha-glycerophosphoric acid
  • Succinic Acid