Abstract
The G protein-coupled receptor (GPCR) opsin is a phospholipid scramblase that facilitates rapid transbilayer phospholipid exchange in liposomes. The mechanism of opsin-mediated lipid scrambling is not known but it has been proposed that the lipid translocation pathway may lie within the interface created by the two protomers of an opsin dimer. To test this proposal we engineered QUAD opsin, in which four lipid-facing residues in transmembrane helix 4 are substituted with tryptophan. Herein we show that QUAD opsin has scramblase activity similar to that of the wild type protein. However, unlike wild type opsin, QUAD opsin reconstitutes as a monomer into the vesicles in which its scrambling activity is measured. In addition to the QUAD mutant, we have found that some unclassified Retinitis Pigmentosa-linked mutations (V209M and F220C in TM5 and F45L in TM1), which are otherwise functional proteins, also scramble as monomers. These results indicate that opsin dimerization is not a prerequisite for lipid scrambling.
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