Abstract
Time-resolved UV/Visible absorbance of the retinal chromophore of rhodopsin [1] delivers vital information about key GPCR activation steps in a membrane lipid environment [2]. Following rhodopsin photoexcitation, a minimum of four intermediates equilibrate on the millisecond-time scale [2,3]. The first equilibrium is between the protonated Schiff base (PSB) species Meta I480 and the deprotonated SB species Meta IIa and is pH independent. A second equilibrium entails spectrally silent conversion of Meta IIa to the Meta IIb substate. The final step is protonation of Glu134 of the ERY sequence in Meta IIb to yield Meta IIbH+ whose pKa characterizes the acid-base equilibrium [3]. Absorbance measurements on the microsecond-to-hundred millisecond-time scale allowed us to study effects of POPC, DOPC, or DOPC/DOPE mixtures on the first equilibrium constant K1 and on the pKa of the final equilibrium. Results were analyzed by singular-value decomposition and were fit by a linear combination of temperature-dependent basis spectra. Notably an increase in K1 was discovered due to either PE head groups or increased acyl chain unsaturation, whereas little change in pKawas evident. According to the flexible-surface model (FSM) rhodopsin becomes a sensor of negative spontaneous (intrinsic) curvature upon light activation [4]. Competition between the curvature elastic energy and the hydrophobic mismatch at the proteolipid boundary explains the influences of lipid-protein interactions [4]. Both the lipid acyl chains and polar head groups affect the Meta I-Meta II transition, revealing how protein energetics are affected by material properties of the lipid bilayer.[1] A.V. Struts et al. (2011) Nature Struct. Mol. Biol. 18, 392-394.[2] J. Epps et al. (2006) Photochem. Photobiol. 82, 1436-1441.[3] M. Mahalingam et al. (2008) PNAS 105 17795-17800.[4] A. V. Botelho et al. (2006) Biophys. J. 91, 4464-4477.
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