Phosphatidylethanolamine enhances rhodopsin photoactivation and transducin binding in a solid supported lipid bilayer as determined using plasmon-waveguide resonance spectroscopy - PubMed (original) (raw)
Phosphatidylethanolamine enhances rhodopsin photoactivation and transducin binding in a solid supported lipid bilayer as determined using plasmon-waveguide resonance spectroscopy
Isabel D Alves et al. Biophys J. 2005 Jan.
Erratum in
- Biophys J. 2006 Jan 15;90(2):709
Abstract
Flash photolysis studies have shown that the membrane lipid environment strongly influences the ability of rhodopsin to form the key metarhodopsin II intermediate. Here we have used plasmon-waveguide resonance (PWR) spectroscopy, an optical method sensitive to both mass and conformation, to probe the effects of lipid composition on conformational changes of rhodopsin induced by light and due to binding and activation of transducin (G(t)). Octylglucoside-solubilized rhodopsin was incorporated by detergent dilution into solid-supported bilayers composed either of egg phosphatidylcholine or various mixtures of a nonlamellar-forming lipid (dioleoylphosphatidylethanolamine; DOPE) together with a lamellar-forming lipid (dioleoylphosphatidylcholine; DOPC). Light-induced proteolipid conformational changes as a function of pH correlated well with previous flash photolysis studies, indicating that the PWR spectral shifts monitored metarhodopsin II formation. The magnitude of these effects, and hence the extent of the conformational transition, was found to be proportional to the DOPE content. Our data are consistent with previous suggestions that lipids having a negative spontaneous curvature favor elongation of rhodopsin during the activation process. In addition, measurements of the G(t)/rhodopsin interaction in a DOPC/DOPE (25:75) bilayer at pH 5 demonstrated that light activation increased the affinity for G(t) from 64 nM to 0.7 nM, whereas G(t) affinity for dark-adapted rhodopsin was unchanged. By contrast, in DOPC bilayers the affinity of G(t) for light-activated rhodopsin was only 18 nM at pH 5. Moreover exchange of GDP for GTP gamma S was also monitored by PWR spectroscopy. Only the light-activated receptor was able to induce this exchange which was unaffected by DOPE incorporation. These findings demonstrate that nonbilayer-forming lipids can alter functionally linked conformational changes of G-protein-coupled receptors in membranes, as well as their interactions with downstream effector proteins.
Figures
FIGURE 1
PWR spectra obtained for egg PC lipid bilayer formation, rhodopsin incorporation in the dark, and light activation, in 10 mM phosphate buffer at pH 5 using _p_-polarized (A) and _s_-polarized (B) light excitation. All spectra were measured with 632.8 nm exciting light. In both A and B, curve 1 represents the buffer spectra before bilayer formation, curve 2 shows PWR spectra obtained after the formation of an egg PC bilayer on the resonator surface, curve 3 shows PWR spectra obtained after addition of an octylglucoside-containing buffer solution of rhodopsin to the aqueous compartment of the PWR cell in the dark (the final rhodopsin concentration in the cell sample compartment was ≈1 _μ_M), and curve 4 shows PWR spectra obtained upon saturating yellow light (λ > 500 nm) activation of rhodopsin.
FIGURE 2
PWR spectra obtained upon light activation of rhodopsin incorporated into an egg PC lipid bilayer in 10 mM phosphate buffer, at pH 7.5, using _p_-polarized (A) and _s_-polarized (B) light excitation. All spectra were measured with 632.8 nm exciting light. (•) PWR spectra obtained after addition in the dark of an octylglucoside-containing buffer solution of rhodopsin to the aqueous compartment of the sample cell containing a preformed bilayer (final rhodopsin concentration in the cell sample compartment was ≈1 _μ_M); (○) PWR spectra obtained upon saturating yellow light (λ > 500 nm) activation of rhodopsin. Solid lines represent the data; symbols are used to distinguish the two spectral curves. Note that the two sets of spectra superimpose on each other within the spectral resolution (1 mdeg) indicating that no rhodopsin photoactivation occurred.
FIGURE 3
Dependence on pH of the changes in the PWR resonance angular position observed in recombinant rhodopsin/egg PC films in 10 mM phosphate buffer upon saturating yellow light irradiation. Squares represent the results obtained with _p_-polarized light and triangles with _s_-polarized light. The solid curve through the data points for both polarizations represents the best fit to the Henderson-Hasselbalch equation with an apparent pKa value of 6.4 ± 0.05.
FIGURE 4
PWR spectra obtained for a rhodopsin/DOPC recombinant film and changes obtained upon yellow light activation at pH 5 using _p_-polarized (A) and _s_-polarized (B) light excitation. All spectra were measured with 632.8 nm exciting light. For both panels, curve 1 represents the PWR spectra obtained after the formation of a DOPC lipid bilayer on the resonator surface, curve 2 shows PWR spectra obtained after addition of an octylglucoside-containing buffer solution of rhodopsin to the aqueous sample compartment in the dark (final rhodopsin concentration in the cell sample compartment was ≈1 _μ_M), and curve 3 shows PWR spectra obtained upon saturating yellow light (λ > 500 nm) activation of rhodopsin.
FIGURE 5
PWR spectra obtained for rhodopsin in DOPC/DOPE (25:75 mol %) recombinant films and changes obtained upon yellow light activation at pH 5 using _p_-polarized (A) and _s_-polarized (B) light excitation. All spectra were measured with 632.8 nm exciting light. For both panels, curve 1 represents the PWR spectra obtained after the formation of a DOPC/DOPE lipid bilayer on the resonator surface, curve 2 shows PWR spectra obtained after addition of an octylglucoside-containing buffer solution of rhodopsin to the aqueous sample compartment in the dark (final rhodopsin concentration in the cell sample compartment was ≈1 _μ_M), and curve 3 shows PWR spectra obtained upon saturating yellow light (λ > 500 nm) activation of rhodopsin.
FIGURE 6
Dependence on pH of the changes in the PWR resonance angular position observed for rhodopsin in DOPC/DOPE (25:75 mol %) recombinant films upon saturating yellow light irradiation. Squares represent the results obtained with _p_-polarized light and triangles those obtained using _s_-polarized light. The solid curve through the data points for both polarizations represents the best fit to the Henderson-Hasselbalch equation with an apparent pKa value of 7.3 ± 0.13.
FIGURE 7
Dependence on the mol %/DOPE of the changes in the PWR resonance angle position observed for rhodopsin incorporated into DOPC/DOPE recombinant films at pH 5 upon saturating yellow light irradiation. Conditions as in Figs. 5 and 6. Squares represent the results obtained with _p_-polarized light and triangles those obtained using _s_-polarized light. The solid line through the data points represents the best linear least-squares fit.
FIGURE 8
Interaction of transducin (Gt) with a control DOPC/DOPE (25:75 mol %) lipid bilayer in the absence of rhodopsin using _p_- (A) and _s_-polarized (B) light excitation. All spectra were measured with 632.8 nm exciting light. In both panels, curve 1 represents the PWR spectra obtained for the DOPC/DOPE lipid bilayer and curve 2 shows PWR spectra after Gt addition to the sample compartment (1 _μ_M was the final Gt concentration in the sample cell).
FIGURE 9
Interaction of transducin (Gt) with dark-adapted rhodopsin in DOPC/DOPE (25:75 mol %) recombinant films at pH 5, and PWR spectral changes obtained upon light activation and GTP_γ_S addition using _p_- (A) and _s_-polarized (B) light excitation. All spectra were measured with 632.8 nm exciting light. In both panels, curve 1 represents the PWR spectra obtained after rhodopsin incorporation into the lipid bilayer (DOPC/DOPE; 25:75 mol %) in the dark, curve 2 shows PWR spectra after Gt addition to the sample compartment in the dark (10 nM is the final Gt concentration in the sample cell), curve 3 shows PWR spectra obtained upon saturating yellow light activation of the rhodopsin-Gt complex, and curve 4 shows the PWR spectra obtained after GTP_γ_S addition to the sample compartment in the dark (final GTP_γ_S concentration in the sample cell was 3 _μ_M).
FIGURE 10
Binding curves for the interaction of Gt with dark-adapted (A) and light-activated (B) rhodopsin in a DOPC/DOPE (25:75 mol %) lipid bilayer at pH 5. Isotherms were obtained by plotting the shifts in the resonance angular minimum of PWR spectra measured after several incremental additions of Gt for _p_- (▪) and _s_-polarized (▴) light. Note the differences in the concentration ranges, and thus the differences in affinity observed in the two panels. The data are fit to a hyperbolic function (solid curves). Dissociation constant values are given in the figure as well as in Table 1.
FIGURE 11
Binding curves for the interaction of GTP_γ_S with the light-activated rhodopsin-Gt complex in DOPC/DOPE (25:75 mol %) films. Isotherms plot the shifts in the resonance angle minimum of the PWR spectra obtained after addition of aliquots of GTP_γ_S for _p_- (▪) and _s_-polarized (▴) light. Solid curves correspond to hyperbolic fits to the data; dissociation constant values are given in Table 1.
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References
- Altenbach, C., K. Cai, H. G. Khorana, and W. L. Hubbell. 1999a. Structural features and light-dependent changes in the sequence 306–322 extending from helix VII to the palmitoylation sites in rhodopsin: a site-directed spin-labeling study. Biochemistry. 38:7931–7937. - PubMed
- Altenbach, C., K. Cai, J. Klein-Seetharaman, H. G. Khorana, and W. L. Hubbell. 2001a. Structure and function in rhodopsin: mapping light-dependent changes in distance between residue 65 in helix TM1 and residues in the sequence 306–319 at the cytoplasmic end of helix TM7 and in helix H8. Biochemistry. 40:15483–15492. - PubMed
- Altenbach, C., J. Klein-Seetharaman, K. Cai, H. G. Khorana, and W. L. Hubbell. 2001b. Structure and function in rhodopsin: mapping light-dependent changes in distance between residue 316 in helix 8 and residues in the sequence 60–75, covering the cytoplasmic end of helices TM1 and TM2 and their connection loop CL1. Biochemistry. 40:15493–15500. - PubMed
- Altenbach, C., J. Klein-Seetharaman, J. Hwa, H. G. Khorana, and W. L. Hubbell. 1999b. Structural features and light-dependent changes in the sequence 59–75 connecting helices I and II in rhodopsin: a site-directed spin-labeling study. Biochemistry. 38:7945–7949. - PubMed
- Alves, I. D., K. A. Ciano, V. Boguslavsky, E. Varga, Z. Salamon, H. I. Yamamura, V. J. Hruby, and G. Tollin. 2004. Selectivity, cooperativity and reciprocity in the interactions between the delta opioid receptor, its ligands and G-proteins. J. Biol. Chem. 279:44673–44682. - PubMed
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