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R Sankararamakrishnan

Publications and source records attributed to R Sankararamakrishnan.

At least 19 recordsLinked to original sources
PubMed1996

Molecular dynamics simulations of isolated transmembrane helices of potassium channels.

In the middle of the S6 helix in voltage-gated potassium channels there is a highly conserved Pro-Val-Pro motif, while the equivalent M2 helix of inward rectifier potassium channels contains a conserved glycine residue in a comparable position. The structural implications of these conserved motifs are of interest given the evidence that S6 and M2 are components of the lining of their respective pores. Multiple sequence alignment and TM helix prediction methods were used to define consensus regions for S6 and M2. Ensembles of 50 structures for each helix were generated by simulated annealing and restrained molecular dynamics. Time-dependent fluctuations of S6 and M2 were investigated by long time scale molecular dynamics simulations on representative members of each ensemble carried out in vacuo in the presence and absence of a hydrophobic potential that mimics a lipid bilayer. The results are discussed in terms of the structural basis of the kink in S6 and M2 and of a putative functional role for flexible helices as "molecular swivels."

Amino Acid Sequence↗
PubMed1996

The pore domain of the nicotinic acetylcholine receptor: molecular modeling, pore dimensions, and electrostatics.

The pore domain of the nicotinic acetylcholine receptor has been modeled as a bundle of five kinked M2 helices. Models were generated via molecular dynamics simulations incorporating restraints derived from 9-A resolution cryoelectron microscopy data (Unwin, 1993; 1995), and from mutagenesis data that identify channel-lining side chains. Thus, these models conform to current experimental data but will require revision as higher resolution data become available. Models of the open and closed states of a homopentameric alpha 7 pore are compared. The minimum radius of the closed-state model is less than 2 A; the minimum radius of the open-state models is approximately 6 A. It is suggested that the presence of "bound" water molecules within the pore may reduce the effective minimum radii below these values by up to approximately 3 A. Poisson-Boltzmann calculations are used to obtain a first approximation to the potential energy of a monovalent cation as it moves along the pore axis. The differences in electrostatic potential energy profiles between the open-state models of alpha 7 and of a mutant of alpha 7 are consistent with the experimentally observed change in ion selectivity from cationic to anionic. Models of the open state of the heteropentameric Torpedo nicotinic acetylcholine receptor pore domain are also described. Relatively small differences in pore radius and electrostatic potential energy profiles are seen when the Torpedo and alpha 7 models are compared.

Amino Acid Sequence↗
PubMed1996

Solvation, water permeation, and ionic selectivity of a putative model for the pore region of the voltage-gated sodium channel.

This paper describes a molecular dynamics and molecular mechanics study of the solvation and selectivity of the narrow pore and vestibule region of a model-built structure for the voltage-gated sodium channel. The particular structure used was one proposed by Guy and Durell. However, many of the features we saw would likely be shared with other possible models for this channel, such as the one proposed by Lipkind and Fozzard. It was found that the water mobility was reduced in the channel and the water orientations were significantly ordered by the channel environment. Water mobility depended on protein mobility; in a computer experiment in which the protein was artificially frozen, channel water at 300 degrees K was immobilized. Water motions were defined in significant part by a series of discrete moves from one pattern of hydrogen bonding with particular amino acids to another. However, there are so many different hydrogen bonding patterns that a description of the motion in terms of transitions among a small number of discrete states is not appropriate. In the model whose solvation we explored, several charged residues seem to play a particularly significant role in determining solvation and water motions. Based on energy minimization studies, the structure clearly shows selectivity for univalent cations over anions.

Amino Acid Sequence↗
PubMed1996

Water in channel-like cavities: structure and dynamics.

Ion channels contain narrow columns of water molecules. It is of interest to compare the structure and dynamics of such intrapore water with those of the bulk solvent. Molecular dynamics simulations of modified TIP3P water molecules confined within channel-like cavities have been performed and the orientation and dynamics of the water molecules analyzed. Channels were modeled as cylindrical cavities with lengths ranging from 15 to 60 A and radii from 3 to 12 A. At the end of the molecular dynamics simulations water molecules were observed to be ordered into approximately concentric cylindrical shells. The waters of the outermost shell were oriented such that their dipoles were on average perpendicular to the normal of the wall of the cavity. Water dynamics were analyzed in terms of self-diffusion coefficients and rotational reorientation rates. For cavities of radii 3 and 6 A, water mobility was reduced relative to that of simulated bulk water. For 9- and 12-A radii confined water molecules exhibited mobilities comparable with that of the bulk solvent. If water molecules were confined within an hourglass-shaped cavity (with a central radius of 3 A increasing to 12 A at either end) a gradient of water mobility was observed along the cavity axis. Thus, water within simple models of transbilayer channels exhibits perturbations of structure and dynamics relative to bulk water. In particular the reduction of rotational reorientation rate is expected to alter the local dielectric constant within a transbilayer pore.

Biophysical Phenomena↗
PubMed1996

Molecular dynamics simulations of water within models of ion channels.

The transbilayer pores formed by ion channel proteins contain extended columns of water molecules. The dynamic properties of such waters have been suggested to differ from those of water in its bulk state. Molecular dynamics simulations of ion channel models solvated within and at the mouths of their pores are used to investigate the dynamics and structure of intra-pore water. Three classes of channel model are investigated: a) parallel bundles of hydrophobic (Ala20) alpha-helices; b) eight-stranded hydrophobic (Ala10) antiparallel beta-barrels; and c) parallel bundles of amphipathic alpha-helices (namely, delta-toxin, alamethicin, and nicotinic acetylcholine receptor M2 helix). The self-diffusion coefficients of water molecules within the pores are reduced significantly relative to bulk water in all of the models. Water rotational reorientation rates are also reduced within the pores, particularly in those pores formed by alpha-helix bundles. In the narrowest pore (that of the Ala20 pentameric helix bundle) self-diffusion coefficients and reorientation rates of intra-pore waters are reduced by approximately an order of magnitude relative to bulk solvent. In Ala20 helix bundles the water dipoles orient antiparallel to the helix dipoles. Such dipole/dipole interaction between water and pore may explain how water-filled ion channels may be formed by hydrophobic helices. In the bundles of amphipathic helices the orientation of water dipoles is modulated by the presence of charged side chains. No preferential orientation of water dipoles relative to the pore axis is observed in the hydrophobic beta-barrel models.

Alamethicin↗
PubMed1995

Water-mediated conformational transitions in nicotinic receptor M2 helix bundles: a molecular dynamics study.

The ion channel of the nicotinic acetylcholine receptor is a water-filled pore formed by five M2 helix segments, one from each subunit. Molecular dynamics simulations on bundles of five M2 alpha 7 helices surrounding a central column of water and with caps of water molecules at either end of the pore have been used to explore the effects of intrapore water on helix packing. Interactions of water molecules with the N-terminal polar sidechains lead to a conformational transition from right- to left-handed supercoils during these stimulations. These studies reveal that the pore formed by the bundle of M2 helices is flexible. A structural role is proposed for water molecules in determining the geometry of bundles of isolated pore-forming helices.

Amino Acid Sequence↗
PubMed1995

Modelling packing interactions in parallel helix bundles: pentameric bundles of nicotinic receptor M2 helices.

The transbilayer pore of the nicotinic acetylcholine receptor (nAChR) is formed by a pentameric bundle of M2 helices. Models of pentameric bundles of M2 helices have been generated using simulated annealing via restrained molecular dynamics. The influence of: (a) the initial C alpha template; and (b) screening of sidechain electrostatic interactions on the geometry of the resultant M2 helix bundles is explored. Parallel M2 helices, in the absence of sidechain electrostatic interactions, pack in accordance with simple ridges-in-grooves considerations. This results in a helix crossing angle of ca. +12 degrees, corresponding to a left-handed coiled coil structure for the bundle as a whole. Tilting of M2 helices away from the central pore axis at their C-termini and/or inclusion of sidechain electrostatic interactions may perturb such ridges-in-grooves packing. In the most extreme cases right-handed coiled coils are formed. An interplay between inter-helix H-bonding and helix bundle geometry is revealed. The effects of changes in electrostatic screening on the dimensions of the pore mouth are described and the significance of these changes in the context of models for the nAChR pore domain is discussed.

Amino Acid Sequence↗
PubMed1995

Packing interactions of Aib-containing helices: molecular modeling of parallel dimers of simple hydrophobic helices and of alamethicin.

alpha-Aminoisobutyric acid (Aib) is a helicogenic alpha, alpha-dimethyl amino acid found in channel-forming peptaibols such as alamethicin. Possible effects of Aib on helix-helix packing are analyzed. Simulated annealing via restrained molecular dynamics is used to generate ensembles of approximately parallel helix dimers. Analysis of variations in geometrical and energetic parameters within ensembles defines how tightly a pair of helices interact. Simple hydrophobic helix dimers are compared: Ala20, Leu20, Aib20, and P20, the latter a simple channel-forming peptide [G. Menestrina, K.P. Voges, G. Jung, and G. Boheim (1986) Journal of Membrane Biology, Vol. 93, pp. 111-132]. Ala20 and Leu20 dimers exhibit well-defined ridges-in-grooves packing with helix crossing angles (omega) of the order of +20 degrees. Aib20 alpha-helix dimers are much more loosely packed, as evidenced by a wide range of omega values and small helix-helix interaction energies. However, when in a 3(10) conformation Aib20 helices pack in three well-defined parallel modes, with omega ca. -15 degrees, +5 degrees, and 10 degrees. Comparison of helix-helix interaction energies suggests that dimerization may favor the 3(10) conformation. P20, with 8 Aib residues, also shows looser packing of alpha-helices. The results of these studies of hydrophobic helix dimers are analyzed in the context of the ridges-in-grooves packing model. Simulations are extended to dimers of alamethicin, and of an alamethicin derivative in which all Aib residues are replaced by Leu. This substitution has little effect on helix-helix packing. Rather, such interactions appear to be sensitive to interactions between polar side chains. Overall, the results suggest that Aib may modulate the packing of simple hydrophobic helices, in favor of looser interactions. For more complex amphipathic helices, interactions between polar side chains may be more critical.

Alamethicin↗
PubMed1995

Structural features of isolated M2 helices of nicotinic receptors. Simulated annealing via molecular dynamics studies.

The nicotinic acetylcholine receptor is an integral membrane protein and a ligand-gated cation channel. It has stoichiometry alpha 2 beta gamma delta, the subunits arranged symmetrically around an approximate five-fold axis. Five M2 helices, one from each subunit, form a parallel helix bundle surrounding a central pore. Simulated annealing via restrained molecular dynamics (SA/MD) has been employed to generate ensembles of isolated M2 transmembrane helices. Four ensembles of two different M2 helix sequences, M2 delta and M2 gamma, have been generated by SA/MD. The ensembles differed in their treatment of electrostatic interactions. Analysis of the simulated structures showed that intra-helical H-bonds were more strongly conserved in the C-terminal (and more hydrophobic) segment of M2 helices. Conformations of polar sidechains have been analyzed, placing particular emphasis on EK (and QK) pairs at the N-termini of M2 delta (and M2 gamma) helices. Conformations of EK sidechain pairs were obtained for the high resolution structures in the protein database in order to guide our analysis of simulated structures. Serine and threonine sidechain conformations in the M2 models also have been determined. Implications of studies of isolated M2 helices for models of the intact pore region of the nicotinic receptor are discussed.

Amino Acid Sequence↗
PubMed1995

Seven-helix bundles: molecular modeling via restrained molecular dynamics.

Simulated annealing via restrained molecular dynamics (SA/MD) has been used to model compact bundles of seven approximately (anti)parallel alpha-helices. Seven such helix bundles occur, e.g., in bacteriorhodopsin, in rhodopsin, and in the channel-forming N-terminal domain of Bacillus thuringiensis delta-endotoxin. Two classes of model are considered: (a) those consisting of seven Ala20 peptide chains; and (b) those containing a single polypeptide chain, made up of seven Ala20 helices linked by GlyN interhelix loops (where N = 5 or 10). Three different starting C alpha templates for SA/MD are used, in which the seven helices are arranged (a) on a left-handed circular template, (b) on a bacteriorhodopsin-like template, or (c) on a zig-zag template. The ensembles of models generated by SA/MD are analyzed in terms of their geometry and energetics, and the most stable structures from each ensemble are examined in greater detail. Structures resembling bacteriorhodopsin and structures resembling delta-endotoxin are both represented among the most stable structures. delta-Endotoxin-like structures arise from both circular and bacteriorhodopsin-like C alpha templates. A third helix-packing mode occurs several times among the stable structures, regardless of the C alpha template and of the presence or absence of interhelix loops. It is characterized by a "4 + 1" core, in which four helices form a distorted left-handed supercoil around a central, buried helix. The remaining two helices pack onto the outside of the core. This packing mode is comparable with that proposed for rhodopsin on the basis of two-dimensional electron crystallographic and sequence analysis studies.

Amino Acid Sequence↗
PubMed1995

Modelling membrane proteins using structural restraints.

Here we present a procedure for modelling membrane proteins which employs molecular dynamics simulations incorporating target restraints derived from low-resolution structures alongside distance restraints derived from mutagenesis data. The application of the modelling procedure to the closed conformation of the pore domain of the nicotinic acetylcholine receptor is described. This domain is formed by a parallel bundle of five M2 helices. Each M2 helix is kinked due to cumulative distortions of backbone (phi, psi) values. The central region of M2 may adopt a more distorted conformation. This would enable a ring of conserved leucine residues (one from each M2 helix) to pack together, occluding the central pore and thus preventing ion permeation. Molecular dynamics simulations on isolated helices that kink formation is not an inherent property of M2.

Amino Acid Sequence↗
PubMed1994

Kinked structures of isolated nicotinic receptor M2 helices: a molecular dynamics study.

The pore-lining M2 helix of the nicotinic acetylcholine receptor exhibits a pronounced kink when the corresponding ion channel is in a closed conformation [N. Unwin (1993) Journal of Molecular Biology, Vol. 229, pp. 1101-1124]. We have performed molecular dynamics simulations of isolated 22-residue M2 helices in order to identify a possible molecular origin of this kink. In order to sample a wide range of conformational space, a simulated annealing protocol was used to generate five initial M2 helix structures, each of which was subsequently used as the basis of 300 ps MD simulations. Two helix sequences (M2 alpha and M2 delta) were studied in this manner, resulting in a total of ten 300 ps trajectories. Kinked helices present in the trajectories were identified and energy minimized to yield a total of five different stable kinked structures. For comparison, a similar molecular dynamics simulation of a Leu23 helix yielded no stable kinked structures. In four of the five kinked helices, the kink was stabilized by H bonds between the helix backbone and polar side-chain atoms. Comparison with data from the literature on site-directed mutagenesis of M2 residues suggests that such polar side-chain to main-chain H bonds may also contribute to kinking of M2 helices in the intact channel protein.

Amino Acid Sequence↗
PubMed1994

Parallel helix bundles and ion channels: molecular modeling via simulated annealing and restrained molecular dynamics.

A parallel bundle of transmembrane (TM) alpha-helices surrounding a central pore is present in several classes of ion channel, including the nicotinic acetylcholine receptor (nAChR). We have modeled bundles of hydrophobic and of amphipathic helices using simulated annealing via restrained molecular dynamics. Bundles of Ala20 helices, with N = 4, 5, or 6 helices/bundle were generated. For all three N values the helices formed left-handed coiled coils, with pitches ranging from 160 A (N = 4) to 240 A (N = 6). Pore radius profiles revealed constrictions at residues 3, 6, 10, 13, and 17. A left-handed coiled coil and a similar pattern of pore constrictions were observed for N = 5 bundles of Leu20. In contrast, N = 5 bundles of Ile20 formed right-handed coiled coils, reflecting loosened packing of helices containing beta-branched side chains. Bundles formed by each of two classes of amphipathic helices were examined: (a) M2a, M2b, and M2c derived from sequences of M2 helices of nAChR; and (b) (LSSLLSL)3, a synthetic channel-forming peptide. Both classes of amphipathic helix formed left-handed coiled coils. For (LSSLLSL)3 the pitch of the coil increased as N increased from 4 to 6. The M2c N = 5 helix bundle is discussed in the context of possible models of the pore domain of nAChR.

Amino Acid Sequence↗
PubMed1993

Characterization of proline-containing alpha-helix (helix F model of bacteriorhodopsin) by molecular dynamics studies.

Many of the bilayer spanning segments of membrane transport proteins contain proline residues, and most of them are believed to occur in alpha-helical form. A proline residue in the middle of an alpha-helix is known to produce a bend in the helix, and recent studies have focused on characterizing such a bend at atomic level. In the present case, molecular dynamics (MD) studies are carried out on helix F model of bacteriorhodopsin (BR) Ace-(Ala)7-Trp-(Ala)2-Tyr-Pro-(Ala)2-Trp- (Ala)8-NHMe and compared with Ace-(Ala)7-Trp-(Ala)2-Tyr-(Ala)3-Trp-(Ala)8-NHMe in which the proline is replaced by alanine. The bend in the helix is characterized by structural parameters such as kink angle (alpha), wobble angle (theta), virtual torsion angle (rho), and the hydrogen bond distance d (Op-3 ... Np+1). The average values and the flexibility involved in these parameters are evaluated. The correlation among the bend related parameters are estimated. The equilibrium side chain orientations of tryptophan and tyrosine residues are discussed and compared with those found in the recently proposed model of bacteriorhodopsin. Finally, a detailed characterization of the bend in terms of secondary structures such as alpha I, alpha II and goniometric helices are discussed, which can be useful in the interpretation of the experimental results on the secondary structures of membrane proteins involving the proline residue.

Amino Acid Sequence↗