Abstract
Mitochondrial inner membrane uncoupling proteins (UCPs) facilitate transmembrane (TM) proton flux and consequently reduce the membrane potential and ATP production. It has been proposed that the three neuronal human UCPs (UCP2, UCP4 and UCP5) in the central nervous system (CNS) play significant roles in reducing cellular oxidative stress. However, the structure and ion transport mechanism of these proteins remain relatively unexplored. Recently, we reported a novel expression system for obtaining functionally folded UCP1in bacterial membranes and applied this system to obtain highly pure neuronal UCPs in high yields. In the present study, we report on the structure and function of the three neuronal UCP homologues. Reconstituted neuronal UCPs were dominantly helical in lipid membranes and transported protons in the presence of physiologically-relevant fatty acid (FA) activators. Under similar conditions, all neuronal UCPs also exhibited chloride transport activities that were partially inhibited by FAs. CD, fluorescence and MS measurements and semi-native gel electrophoresis collectively suggest that the reconstituted proteins self-associate in the lipid membranes. Based on SDS titration experiments and other evidence, a general molecular model for the monomeric, dimeric and tetrameric functional forms of UCPs in lipid membranes is proposed. In addition to their shared structural and ion transport features, neuronal UCPs differ in their conformations and proton transport activities (and possibly mechanism) in the presence of different FA activators. The differences in FA-activated UCP-mediated proton transport could serve as an essential factor in understanding and differentiating the physiological roles of UCP homologues in the CNS.
Highlights
Located in the inner mitochondrial membrane (IMM), uncoupling proteins (UCPs) dissipate the proton electrochemical gradient across the membrane, resulting in the reduction of ATP production through ATP synthase
Using planar lipid bilayer systems, it was shown that arachidonic acid (AA) is a strong fatty acid (FA) activator for the proton transport activity of UCP2 [35]
To further examine the ion transport mechanism of UCPs, we examined the effects of FAs (PA and AA) on the chloride transport activity of neuronal UCPs in proteoliposomes
Summary
Located in the inner mitochondrial membrane (IMM), uncoupling proteins (UCPs) dissipate the proton electrochemical gradient across the membrane, resulting in the reduction of ATP production through ATP synthase. Compared with the prototypic UCP1, the amino acid sequence identities of UCPs 2, 4, 5 are 59 %, 34 % and 30 % respectively [8,9,10,11]. Despite these relatively low sequence identities to UCP1, both UCP4 and UCP5 possess comparable helical conformations and proton transport c 2015 Authors.
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