Key Differences in Molecular Transport Mechanisms of Uncoupling Proteins

Abstract

Mitochondrial membrane uncoupling protein 1 (UCP1) facilitates proton leak to support non-shivering thermogenesis in brown adipose tissue. The transport function of other UCPs is controversially discussed. It was proposed that in addition to protons, other substrates can be transported. Moreover, the molecular mechanism of UCP regulation is insufficiently understood, although it is generally accepted that its transport is regulated by free fatty acids (FFA) and purine nucleotides (PN). Here we tested the hypothesis that regulation differs between members of UCP family leading to differences in their functions [1]. We evaluated binding forces and the degree of inhibition by PNs, comparing the data obtained using recognition force microscopy and electrophysiological measurements of recombinant proteins reconstituted in planar bilayer membranes [2]. We reveal that, in contrast to UCP1, other UCPs can be fully inhibited by all PNs, as KD increases with a decrease in phosphorylation. Furthermore, three arginines (R84, R183, R277) in the PN-binding pocket are involved in UCP1 inhibition to different extents. This result disagrees with previously proposed mechanisms, suggesting that only R277 is responsible for 100% inhibition. Moreover, FFAs can compete with all PNs bound to UCP1, but only with triphosphate-PNs bound to UCP3. Our results demonstrate the different regulation across a family of highly homologous uncoupling proteins, which, in the case of UCP1 and UCP3, are even expressed in the same tissue. We anticipate that the differences in the molecular mechanism of UCPs can be useful in understanding their physiological functions.[1] Rupprecht A, Sokolenko EA, Beck V, Ninnemann O, Jaburek M, et al. (2010) Biophys J; 98: 1503-1511.[2] Zhu R, Rupprecht A, Ebner A, Haselgrubler T, Gruber HJ, Hinterdorfer P, Pohl EE. (2013) J Am Chem Soc; 135:3640-3646.