Fatty acid binding site of mitochondrial uncoupling protein UCP2 as probed by EPR spectroscopy of spin-labeled fatty acids

N. Raju, T. Špaček, J. Ježek, I. M. Caminiti, F. Leinisch, K. Hideg, P. Ježek, Wolfgang E. Trommer

Research output: Article

2 Citations (Scopus)

Abstract

Electron paramagnetic resonance (EPR) spectroscopy of spin-labeled fatty acids was used to investigate their interaction with Escherichia coli-expressed human mitochondrial uncoupling protein UCP2 refolded from inclusion bodies in nonaethylene glycol monododecyl ether (C12E9) micelles. 5-DOXYL-stearic acid and 4-PROXYL-palmitic acid bound to UCP2 exhibited additional clearly separated h+1I, h-1I "immobile" peaks in the low- and high-field region, respectively, separated by 42 and 44 Gauss, and extensively reduced h+1M, h -1M "mobile" peaks, separated by about 30 G, whereas with 7-DOXYL-stearic acid the I and M peaks were smoothed together into one wide peak. Competition of 4-PROXYL-palmitic acid with added palmitic acid, arachidonic acid, and all-cis-8,11,14-eicosatrienoic acid and of 7-DOXYL-stearic acid with arachidonic acid was indicated by the disappearance of the h +1I, h-1I "immobile" peaks, whereas redistribution in micelles without protein was indicated by the rising of the h+1M, h-1M "mobile" peaks. In conclusion, a competition of palmitic, arachidonic, and eicosatrienoic acid within a putative fatty acid binding site was observed for mitochondrial uncoupling protein UCP2. This finding together with the observation of EPR spectra of highly immobilized probes exclusively in the presence of the recombinant UCP2 suggest the existence of a fatty acid binding site on UCP2 which is a prerequisite of the fatty acid cycling mechanism as previously postulated for UCP1.

Original languageEnglish
Pages (from-to)373-383
Number of pages11
JournalApplied Magnetic Resonance
Volume30
Issue number3-4
Publication statusPublished - dec. 1 2006

ASJC Scopus subject areas

  • Atomic and Molecular Physics, and Optics

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    Raju, N., Špaček, T., Ježek, J., Caminiti, I. M., Leinisch, F., Hideg, K., Ježek, P., & Trommer, W. E. (2006). Fatty acid binding site of mitochondrial uncoupling protein UCP2 as probed by EPR spectroscopy of spin-labeled fatty acids. Applied Magnetic Resonance, 30(3-4), 373-383.