Rhodopsin activation affects the environment of specific neighboring phospholipids: An FTIR spectroscopic study

Rhodopsin is a member of a superfamily of G-protein-coupled receptors that transduce signals across membranes. We used Fourier-transform infrared (FTI R) difference spectroscopy to study the interaction between rhodopsin and l ipid bilayer upon receptor activation. A difference band at 1744 cm(-1) (+) /1727 cm(-1) (-) was identified in the FTIR-difference spectrum of rhodopsi n mutant D83N/E122Q in which spectral difference bands arising from the car bonyl stretching frequencies of protonated carboxylic acid groups were remo ved by mutation. As the band was abolished by detergent delipidation, we su ggested that it arose from carbonyl groups of phospholipid fatty acid ester s. Rhodopsin and the D83N/E122Q mutant were reconstituted into various C-13 -labeled 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine vesicles and prob ed. The 1744-cm(-1) (+)/1727 cm(-1) (-) band could be unequivocally assigne d to a change in the lipid ester carbonyl stretch upon receptor activation, with roughly equal contribution from both lipid esters. The band intensity scaled with the amount of rhodopsin but not with the amount of lipid, excl uding the possibility that it was due to the bulk lipid phase. We also excl uded the possibility that the lipid band represents a change in the number of boundary lipids or a general alteration in the boundary lipid environmen t upon formation of metarhodopsin II. Instead, the data suggest that the li pid band represents the change of a specific lipid-receptor interaction tha t is coupled to protein conformational changes.