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Energetics of side-chain partitioning of β-signal residues in unassisted folding of a transmembrane β-barrel protein.
Iyer, BR, Zadafiya, P, Vetal, PV, Mahalakshmi, R
The Journal of biological chemistry. 2017;(29):12351-12365
Abstract
The free energy of water-to-interface amino acid partitioning is a major contributing factor in membrane protein folding and stability. The interface residues at the C terminus of transmembrane β-barrels form the β-signal motif required for assisted β-barrel assembly in vivo but are believed to be less important for β-barrel assembly in vitro Here, we experimentally measured the thermodynamic contribution of all 20 amino acids at the β-signal motif to the unassisted folding of the model β-barrel protein PagP. We obtained the partitioning free energy for all 20 amino acids at the lipid-facing interface (ΔΔG0w,i(φ)) and the protein-facing interface (ΔΔG0w,i(π)) residues and found that hydrophobic amino acids are most favorably transferred to the lipid-facing interface, whereas charged and polar groups display the highest partitioning energy. Furthermore, the change in non-polar surface area correlated directly with the partitioning free energy for the lipid-facing residue and inversely with the protein-facing residue. We also demonstrate that the interface residues of the β-signal motif are vital for in vitro barrel assembly, because they exhibit a side chain-specific energetic contribution determined by the change in nonpolar accessible surface. We further establish that folding cooperativity and hydrophobic collapse are balanced at the membrane interface for optimal stability of the PagP β-barrel scaffold. We conclude that the PagP C-terminal β-signal motif influences the folding cooperativity and stability of the folded β-barrel and that the thermodynamic contributions of the lipid- and protein-facing residues in the transmembrane protein β-signal motif depend on the nature of the amino acid side chain.
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Comparative study of interactions of aliskiren and AT1 receptor antagonists with lipid bilayers.
Sadeghpour, A, Rappolt, M, Ntountaniotis, D, Chatzigeorgiou, P, Viras, K, Megariotis, G, Papadopoulos, MG, Siapi, E, Mali, G, Mavromoustakos, T
Biochimica et biophysica acta. 2015;(4):984-94
Abstract
The renin-angiotensin-aldosterone system (RAAS) plays a key role in the regulation of blood pressure. Renin is the rate limiting enzyme of the RAAS and aliskiren is a highly potent and selective inhibitor of the human renin. Renin is known to be active both in the circulating blood stream as well as locally, when bound to the (pro)-renin receptor ((P)RR). In this study we have investigated a possible mechanism of action of aliskiren, in which its accumulation in the plasma membrane is considered as an essential step for effective inhibition. Aliskiren's interactions with model membranes (cholesterol rich and poor) have been investigated by applying different complementary techniques: differential scanning calorimetry (DSC), Raman spectroscopy, magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy and small- and wide-angle X-ray scattering (SAXS and WAXS). In addition, in silico molecular dynamics (MD) calculations were applied for further confirmation of the experimental data. Aliskiren's thermal effects on the pre- and main transition of dipalmitoyl-phosphatidylcholine (DPPC) membranes as well as its topographical position in the bilayer show striking similarities to those of angiotensin II type 1 receptor (AT1R) antagonists. Moreover, at higher cholesterol concentrations aliskiren gets expelled from the membrane just as it has been recently demonstrated for the angiotensin receptor blocker (ARB) losartan. Thus, we propose that both the AT1R and the (P)RR-bound renin active sites can be efficiently blocked by membrane-bound ARBs and aliskiren when cholesterol rich membrane rafts/caveolae are formed in the vicinity of the receptors.
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3.
Lateral sorting in model membranes by cholesterol-mediated hydrophobic matching.
Kaiser, HJ, Orłowski, A, Róg, T, Nyholm, TK, Chai, W, Feizi, T, Lingwood, D, Vattulainen, I, Simons, K
Proceedings of the National Academy of Sciences of the United States of America. 2011;(40):16628-33
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Abstract
Theoretical studies predict hydrophobic matching between transmembrane domains of proteins and bilayer lipids to be a physical mechanism by which membranes laterally self-organize. We now experimentally study the direct consequences of mismatching of transmembrane peptides of different length with bilayers of different thicknesses at the molecular level. In both model membranes and simulations we show that cholesterol critically constrains structural adaptations at the peptide-lipid interface under mismatch. These constraints translate into a sorting potential and lead to selective lateral segregation of peptides and lipids according to their hydrophobic length.
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4.
Tilt and azimuthal angles of a transmembrane peptide: a comparison between molecular dynamics calculations and solid-state NMR data of sarcolipin in lipid membranes.
Shi, L, Cembran, A, Gao, J, Veglia, G
Biophysical journal. 2009;(9):3648-62
Abstract
We report molecular dynamics simulations in the explicit membrane environment of a small membrane-embedded protein, sarcolipin, which regulates the sarcoplasmic reticulum Ca-ATPase activity in both cardiac and skeletal muscle. In its monomeric form, we found that sarcolipin adopts a helical conformation, with a computed average tilt angle of 28 +/- 6 degrees and azymuthal angles of 66 +/- 22 degrees, in reasonable accord with angles determined experimentally (23 +/- 2 degrees and 50 +/- 4 degrees, respectively) using solid-state NMR with separated-local-field experiments. The effects of time and spatial averaging on both (15)N chemical shift anisotropy and (1)H/(15)N dipolar couplings have been analyzed using short-time averages of fast amide out-of-plane motions and following principal component dynamic trajectories. We found that it is possible to reproduce the regular oscillatory patterns observed for the anisotropic NMR parameters (i.e., PISA wheels) employing average amide vectors. This work highlights the role of molecular dynamics simulations as a tool for the analysis and interpretation of solid-state NMR data.
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5.
A comparative molecular dynamics analysis of the amyloid beta-peptide in a lipid bilayer.
Lemkul, JA, Bevan, DR
Archives of biochemistry and biophysics. 2008;(1):54-63
Abstract
Because the amyloid beta-peptide (Abeta) functions as approximately half of the transmembrane domain of the amyloid precursor protein and interaction of Abeta with membranes is proposed to result in neurotoxicity, the association of Abeta with membranes likely is important in the etiology of Alzheimer's disease. Atomic details of the interaction of Abeta with membranes are not accessible with most experimental techniques, but computational methods can provide this information. Here, we present the results of ten 100-ns molecular dynamics (MD) simulations of the 40-residue amyloid beta-peptide (Abeta40) embedded in a dipalmitoylphosphatidylcholine (DPPC) bilayer. The present study examines the effects of insertion depth, protonation state of key residues, and ionic strength on Abeta40 in a DPPC bilayer. In all cases, a portion of the peptide remained embedded in the bilayer. In the case of deeper insertion depth, Abeta40 adopted a near-transmembrane orientation, drawing water molecules into the bilayer to associate with its charged amino acids. In the case of shallower insertion, the most widely-accepted construct, the peptide associated strongly with the membrane-water interface and the phosphatidylcholine headgroups of the bilayer. In most cases, significant disordering of the extracellular segment of the peptide was observed, and the brief appearance of a beta-strand was noted in one case. Our results compare well with a variety of experimental and computational findings. From this study, we conclude that Abeta associated with membranes is dynamic and capable of adopting a number of conformations, each of which may have significance in understanding the progression of Alzheimer's disease.
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Cholesterol surrogates: a comparison of cholesterol and 16:0 ceramide in POPC bilayers.
Pandit, SA, Chiu, SW, Jakobsson, E, Grama, A, Scott, HL
Biophysical journal. 2007;(3):920-7
Abstract
Experimental evidence indicates that, under some circumstances, "surrogate" molecules may play the same role as cholesterol in ordering membrane lipids. The simplest molecule in this class is Ceramide. In this article, we describe atomic-level molecular dynamics simulations designed to shed light on this phenomenon. We run simulations of hydrated phosphoryl-oleoyl phosphatidylcholine (POPC) bilayers containing cholesterol, and containing ceramide, in concentrations ranging from 5% to 33%. We also perform a simulation of a pure POPC bilayer to verify the simulation force fields against experimental structural data for POPC. Our simulation data are in good agreement with experimental data for the partial molecular volumes, areas, form factors, and order parameters. These simulations suggest that ceramide and cholesterol have a very similar effect on the POPC bilayer, although ceramide is less effective in inducing order in the bilayer compared with cholesterol at the same concentrations.
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7.
Stalk phase formation: effects of dehydration and saddle splay modulus.
Kozlovsky, Y, Efrat, A, Siegel, DP, Kozlov, MM
Biophysical journal. 2004;(4):2508-21
Abstract
One of the earliest lipid intermediates forming in the course of membrane fusion is the lipid stalk. Although many aspects of the stalk hypothesis were elaborated theoretically and confirmed by experiments it remained unresolved whether stalk formation is always an energy consuming process or if there are conditions where the stalks are energetically favorable and form spontaneously resulting in an equilibrium stalk phase. Motivated by a recent breakthrough experiments we analyze the physical factors determining the spontaneous stalk formation. We show that this process can be driven by interplay between two factors: the elastic energy of lipid monolayers including a contribution of the saddle splay deformation and the energy of hydration repulsion acting between apposing membranes. We analyze the dependence of stalk formation on the saddle splay (Gaussian) modulus of the lipid monolayers and estimate the values of this modulus based on the experimentally established phase boundary between the lamellar and the stalk phases. We suggest that fusion proteins can induce stalk formation just by bringing the membranes into close contact, and accumulating, at least locally, a sufficiently large energy of the hydration repulsion.
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8.
Area per lipid and acyl length distributions in fluid phosphatidylcholines determined by (2)H NMR spectroscopy.
Petrache, HI, Dodd, SW, Brown, MF
Biophysical journal. 2000;(6):3172-92
Abstract
Deuterium ((2)H) NMR spectroscopy provides detailed information regarding the structural fluctuations of lipid bilayers, including both the equilibrium properties and dynamics. Experimental (2)H NMR measurements for the homologous series of 1, 2-diacyl-sn-glycero-3-phosphocholines with perdeuterated saturated chains (from C12:0 to C18:0) have been performed on randomly oriented, fully hydrated multilamellar samples. For each lipid, the C-D bond order parameters have been calculated from de-Paked (2)H NMR spectra as a function of temperature. The experimental order parameters were analyzed using a mean-torque potential model for the acyl chain segment distributions, and comparison was made with the conventional diamond lattice approach. Statistical mechanical principles were used to relate the measured order parameters to the lipid bilayer structural parameters: the hydrocarbon thickness and the mean interfacial area per lipid. At fixed temperature, the area decreases with increasing acyl length, indicating increased van der Waals attraction for longer lipid chains. However, the main effect of increasing the acyl chain length is on the hydrocarbon thickness rather than on the area per lipid. Expansion coefficients of the structural parameters are reported and interpreted using an empirical free energy function that describes the force balance in fluid bilayers. At the same absolute temperature, the phosphatidylcholine (PC) series exhibits a universal chain packing profile that differs from that of phosphatidylethanolamines (PE). Hence, the lateral packing of phospholipids is more sensitive to the headgroup methylation than to the acyl chain length. A fit to the area per lipid for the PC series using the empirical free energy function shows that the PE area represents a limiting value for the packing of fluid acyl chains.