Interplay between membrane curvature and protein conformational equilibrium investigated by solid-state NMR

Abstract

Many membrane proteins sense and induce membrane curvature for function, but structural information about how proteins modulate their structures to cause membrane curvature is sparse. We review our recent solid-state NMR studies of two virus membrane proteins whose conformational equilibrium is tightly coupled to membrane curvature. The influenza M2 proton channel has a drug-binding site in the transmembrane (TM) pore. Previous chemical shift data indicated that this pore-binding site is lost in an M2 construct that contains the TM domain and a curvature-inducing amphipathic helix. We have now obtained chemical shift perturbation, protein-drug proximity, and drug orientation data that indicate that the pore-binding site is restored when the full cytoplasmic domain is present. This finding indicates that the curvature-inducing amphipathic helix distorts the TM structure to interfere with drug binding, while the cytoplasmic tail attenuates this effect. In the second example, we review our studies of a parainfluenza virus fusion protein that merges the cell membrane and the virus envelope during virus entry. Chemical shifts of two hydrophobic domains of the protein indicate that both domains have membrane-dependent backbone conformations, with the β-strand structure dominating in negative-curvature phosphatidylethanolamine (PE) membranes. P NMR spectra andH-P correlation spectra indicate that the β-strand-rich conformation induces saddle-splay curvature to PE membranes and dehydrates them, thus stabilizing the hemifusion state. These results highlight the indispensable role of solid-state NMR to simultaneously determine membrane protein structures and characterize the membrane curvature in which these protein structures exist.

Influenza A M2 Spans the Membrane Bilayer, Perturbs its Organization and Differentiates the Effect of Amantadine and Spiro[pyrrolidine-2,2'-adamantane] AK13 on Lipids

“…Therefore, the drugs mask the effects of M2TM at 0.5-1.7 ppm in the NMR spectrum. Additionally, it has been reported that Aamt drugs bound to the M2TM pore reduce its conformational plasticity 68 which is in agreement with the observed limited protein's effect on DMPC fluidization. When Amt or AK13 (x=0.08) and M2TM (x=0.06) concentrations were used (Figure 3 right), the spectrum at pH 8.0 appeared similar to that of DMPC + M2TM (x=0.03) (Figure 3 left), but the peaks intensities were increased, suggesting instead a higher fluidization of DMPC acyl chains in this case.…”

“…Therefore, the drugs mask the effects of M2TM at 0.5-1.7 ppm in the NMR spectrum. Additionally, it has been reported that Aamt drugs bound to the M2TM pore reduce its conformational plasticity 68 which is in agreement with the observed limited protein's effect on DMPC fluidization. When Amt or AK13 (x=0.08) and M2TM (x=0.06) concentrations were used (Figure 3 3).…”

Biological Membrane Organization and Cellular Signaling

“…Recent solid-state NMR experiments indicated that, in the influenza M2 proton channel, an amphipathic helix induces membrane curvature, which in turn distorts the TM helices of the protein to interfere with drug binding. 334 The same research group also reported that the backbone conformations of both hydrophobic domains of a parainfluenza virus fusion protein are membrane-dependent, adopting a β-strand predominant structure in negative-curvature membranes. 335 3.2.4.…”

“…One better studied example is protein kinase C (PKC), a key enzyme in cellular signaling cascades, whose activity has been shown to be tightly controlled by curvature stress due to the weakened lipid headgroup interactions induced by PE and cholesterol. , The C1 domain of PKC acts as a sensor of membrane curvature and undergoes conformational rearrangements to allow the kinase to access the site of phosphorylation in response to the change of its local membrane environment. Recent solid-state NMR experiments indicated that, in the influenza M2 proton channel, an amphipathic helix induces membrane curvature, which in turn distorts the TM helices of the protein to interfere with drug binding . The same research group also reported that the backbone conformations of both hydrophobic domains of a parainfluenza virus fusion protein are membrane-dependent, adopting a β-strand predominant structure in negative-curvature membranes …”

Differential membrane curvature induced by distinct protein conformers

“…20,21 This interplay between protein structural domains and membrane curvature is being investigated and different structural domains have been related to differential membrane curvature. 22,23 However, the molecular mechanisms linking protein conformation and membrane curvature are still missing.…”