Toward Understanding the Mechanism of Ion Transport Activity of Neuronal Uncoupling Proteins UCP2, UCP4, and UCP5

Abstract

Neuronal uncoupling proteins (UCP2, UCP4, and UCP5) have crucial roles in the function and protection of the central nervous system (CNS). Extensive biochemical studies of UCP2 have provided ample evidence of its participation in proton and anion transport. To date, functional studies of UCP4 and UCP5 are scarce. In this study, we show for the first time that, despite a low level of amino acid sequence identity with the previously characterized UCPs (UCP1-UCP3), UCP4 and UCP5 share their functional properties. Recombinantly expressed in Escherichia coli, UCP2, UCP4, and UCP5 were isolated and reconstituted into liposome systems, where their conformations and ion (proton and chloride) transport properties were examined. All three neuronal UCPs are able to transport protons across lipid membranes with characteristics similar to those of the archetypal protein UCP1, which is activated by fatty acids and inhibited by purine nucleotides. Neuronal UCPs also exhibit transmembrane chloride transport activity. Circular dichroism spectroscopy shows that these three transporters exist in different conformations. In addition, their structures and functions are differentially modulated by the mitochondrial lipid cardiolipin. In total, this study supports the existence of general conformational and ion transport features in neuronal UCPs. On the other hand, it also emphasizes the subtle structural and functional differences between UCPs that could distinguish their physiological roles. Differentiation between structure-function relationships of neuronal UCPs is essential for understanding their physiological functions in the CNS.

Expression, Folding, and Proton Transport Activity of Human Uncoupling Protein-1 (UCP1) in Lipid Membranes

“…Given its localization in the IMM, interaction of CL with UCP1 and its influence on this protein structure and function would not be surprising. In a previous study we reported that the proton transport rates of UCP2 and UCP5 in lipid vesicles were increased in the presence of CL (17).…”

“…Recombinant protein expression in bacterial systems is an efficient and inexpensive method for protein production. However, refolding of UCP1 from inclusion bodies could only yield partially folded proteins in mild detergents and liposomes (6,17,19). The recent new development of bacterial strains, fusion proteins, and alternative expression methods provide other possibilities for recombinant UCP1 to be targeted toward the inner membrane of Escherichia coli (20 -22).…”

“…Reconstitution of UCP1 into Liposomes-The reconstitution of purified UCP1 into liposomes for spectroscopic and transport studies was described previously (17). Briefly, lipids were dissolved in chloroform, dried overnight under a vacuum, and rehydrated in the desired buffer.…”

Superresolution microscopy reveals spatial separation of UCP4 and F 0 F 1 -ATP synthase in neuronal mitochondria

“…One of the widely discussed functions for UCP4-another member of the same subfamily that is localized in neurons and neurosensory cells (6)(7)(8)(9)-is the regulation of ROS by decreasing the pmf (10,11). Although there is no unambiguous evidence revealing the exact UCP4 function, it was shown that UCP4 transports protons similar to UCP1 (12). It is, therefore, assumed that UCP4 and other UCPs possibly compete for protons with other proton-consuming proteins, including ATP synthase, but this phenomenon has not yet been studied in detail (13).…”

UCP2 expression may represent a predictive marker of neoadjuvant chemotherapy effectiveness for locally advanced uterine cervical cancer

“…UCP2 and UCP3 are activated by superoxide from the mitochondrial inner membrane, and can reduce ROS generation (21). UCP4 and UCP5 are specifically localized to the brain, and associated with reducing the mitochondrial membrane potential (22).…”