Flipping for Opsin

Science Signaling  01 Feb 2011:
Vol. 4, Issue 158, pp. ec30
DOI: 10.1126/scisignal.4158ec30

Phospholipid flippases are membrane proteins that rapidly transport (flip) lipids from one side of the bilayer to the other. Different flippases function in different membrane-bound compartments and are distinguished by their ability to catalyze unidirectional or bidirectional flipping and in their requirement (or lack thereof) for ATP. Although unidirectional ATP-dependent flippases have been identified, bidirectional ATP-independent flippases—including those found in photoreceptor discs—have not. Menon et al. determined that unilamellar proteoliposomal vesicles containing solubilized bovine disc membrane proteins, phosphatidylcholine, and NBD-PL (a fluorescent phospholipid reporter that loses fluorescence when reduced by dithionite) underwent a greater decrease in fluorescence in response to external dithionite than did protein-free liposomes. Thus, the fluorescence of protein-free liposomes rapidly decreased by about 50% (consistent with localization of half the NBD-PL to the outer leaflet), whereas that of proteoliposomes decreased about 75% (consistent with translocation of NBD-PL by flippases). Similarly, a greater fraction of NBD-PL was available for extraction by albumin in proteoliposomes than in protein-free liposomes. Analysis of the data indicated that 61 to 69% of the reconstituted proteins acted as flippases, identifying rhodopsin (representing about 85% of disc protein content and far more abundant than any other disc protein) as the most likely candidate. Flippase activity was unaffected by light, suggesting that the apoprotein opsin functioned as a flippase. Indeed, human opsin purified from human embryonic kidney (HEK) 293 cells (in which it was heterologously expressed) showed flippase activity independent of the chromophore 11-cis-retinal or any disc-specific cofactors. Opsin is a member of the family of G protein–coupled receptors (GPCRs), and the β1-adrenergic receptor—another GPCR—also showed flippase activity, suggesting the intriguing possibility that other members of this large family of signaling proteins may also function as phospholipid flippases.

I. Menon, T. Huber, S. Sanyal, S. Banerjee, P. Barré, S. Canis, J. D. Warren, J. Hwa, T. P. Sakmar, A. K. Menon, Opsin is a phospholipid flippase. Curr. Biol. 21, 149–153 (2011). [PubMed]