Exploring models of the influenza A M2 channel: MD simulations in a phospholipid bilayer

The M2 protein of influenza A virus forms homotetrameric helix bundles, which function as proton-selective channels. The native form of the protein is 97 residues long, although peptides representing the transmembrane section display ion channel activity, which (like the native channel) is blocked by the antiviral drug amantadine. As a small ion channel, M2 may provide useful insights into more complex channel systems. Models of tetrameric bundles of helices containing either 18 or 22 residues have been simulated while embedded in a fully hydrated 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphatidylcholine bilayer. Several different starting models have been used. These suggest that the simulation results, at least on a nanosecond time scale, are sensitive to the exact starting structure. Electrostatics calculations carried out on a ring of four ionizable aspartate residues at the N-terminal mouth of the channel suggest that at any one time, only one will be in a charged state. Helix bundle models were mostly stable over the duration of the simulation, and their helices remained tilted relative to the bilayer normal. The M2 helix bundles form closed channels that undergo breathing motions, alternating between a tetramer and a dimer-of-dimers structure. Under these conditions either the channel forms a pocket of trapped waters or it contains a column of waters broken predominantly at the C-terminal mouth of the pore. These waters exhibit restricted motion in the pore and are effectively "frozen" in a way similar to those seen in previous simulations of a proton channel formed by a four-helix bundle of a synthetic leucine-serine peptide (, Biophys. J. 77:2400-2410).     

Conformational dynamics of M2 helices in KirBac channels: helix flexibility in relation to gating via molecular dynamics simulations

KirBac1.1 and 3.1 are bacterial homologues of mammalian inward rectifier K channels. We have performed extended molecular dynamics simulations (five simulations, each of >20 ns duration) of the transmembrane domain of KirBac in two membrane environments, a palmitoyl oleoyl phosphatidylcholine bilayer and an octane slab. Analysis of these simulations has focused on the conformational dynamics of the pore-lining M2 helices, which form the cytoplasmic hydrophobic gate of the channel. Principal components analysis reveals bending of M2, with a molecular hinge at the conserved glycine (Gly134 in KirBac1.1, Gly120 in KirBac3.1). More detailed analysis reveals a dimer-of-dimers type motion. The first two eigenvectors describing the motions of M2 correspond to helix kink and swivel motions. The conformational flexibility of M2 seen in these simulations correlates with differences in M2 conformation between that seen in the X-ray structures of closed channels (KcsA and KirBac) in which the helix is undistorted, and in open channels (e.g. MthK) in which the M2 helix is kinked. Thus, the simulations, albeit on a time scale substantially shorter than that required for channel gating, suggest a gating model in which the intrinsic flexibility of M2 about a molecular hinge is coupled to conformational transitions of an intracellular 'gatekeeper' domain, the latter changing conformation in response to ligand binding.     

Computer simulation of ion channel gating: the M(2) channel of influenza A virus in a lipid bilayer

The transmembrane fragment of the influenza virus M(2) protein forms a homotetrameric channel that transports protons. In this paper, we use molecular dynamics simulations to help elucidate the mechanism of channel gating by four histidines that occlude the channel lumen in the closed state. We test two competing hypotheses. In the "shuttle" mechanism, the delta nitrogen atom on the extracellular side of one histidine is protonated by the incoming proton, and, subsequently, the proton on the epsilon nitrogen atom is released on the opposite side. In the "water-wire" mechanism, the gate opens because of electrostatic repulsion between four simultaneously biprotonated histidines. This allows for proton transport along the water wire that penetrates the gate. For each system, composed of the channel embedded in a hydrated phospholipid bilayer, a 1.3-ns trajectory was obtained. It is found that the states involved in the shuttle mechanism, which contain either single-protonated histidines or a mixture of single-protonated histidines plus one biprotonated residue, are stable during the simulations. Furthermore, the orientations and dynamics of water molecules near the gate are conducive to proton transfer. In contrast, the fully biprotonated state is not stable. Additional simulations show that if only two histidines are biprotonated, the channel deforms but the gate remains closed. These results support the shuttle mechanism but not the gate-opening mechanism of proton gating in M(2).     

Molecular dynamics investigation of membrane-bound bundles of the channel-forming transmembrane domain of viral protein U from the human immunodeficiency virus HIV-1

Molecular dynamics (MD) simulations have been carried out on bundles of the channel-forming transmembrane (TM) domain of the viral protein U (VPU(1-27) and VPU(6-27)) from the human immunodeficiency virus (HIV-1). Simulations of hexameric and pentameric bundles of VPU(6-27) in an octane/water membrane mimetic system suggested that the pentamer is the preferred oligomer. Accordingly, an unconstrained pentameric helix bundle of VPU(1-27) was then placed in a hydrated palmitoyl-oleyl-3-n-glycero-phosphatidylethanolamine (POPE) lipid bilayer and its structural properties calculated from a 3-ns MD run. Some water molecules, initially inside the channel lumen, were expelled halfway through the simulation and the bundle adopted a conical structure reminiscent of previous MD results obtained for VPU(6-27) in an octane/water system. The pore constriction generated may correspond to a closed state of the channel and underlies the relocation of the W residue toward the pore lumen. The relative positions of the helices with respect to the bilayer and their interactions with the lipids are discussed. The observed structure is stabilized via specific interactions between the VPU helices and the carbonyl oxygen atoms of the lipid molecules, particularly at the Q and S residues.     

Electrostatics and the ion selectivity of ligand-gated channels.

The nicotinic acetylcholine receptor (nAChR) is a cation-selective ion channel that opens in response to acetylcholine binding. The related glycine receptor (GlyR) is anion selective. The pore-lining domain of each protein may be modeled as a bundle of five parallel M2 helices. Models of the pore-lining domains of homopentameric nAChR and GlyR have been used in continuum electrostatics calculations to probe the origins of ion selectivity. Calculated pKA values suggest that "rings" of acidic or basic side chains at the mouths of the nAChR or GlyR M2 helix bundles, respectively, may not be fully ionized. In particular, for the nAChR the ring of glutamate side chains at the extracellular mouth of the pore is predicted to be largely protonated at neutral pH, whereas those glutamate side chains in the intracellular and intermediate rings (at the opposite mouth of the pore) are predicted to be fully ionized. Inclusion of the other domains of each protein represented as an irregular cylindrical tube in which the M2 bundles are embedded suggests that both the M2 helices and the extramembrane domains play significant roles in determining ion selectivity.     

Distribution and dynamics of adamantanes in a lipid bilayer

The adamantanes are a class of compounds that have found use in the treatment of influenza A and Parkinson's disease, among others. The mode of action for influenza A is based on the adamantanes’ interaction with the transmembrane M2 channel, whereas the treatment of Parkinson's disease is thought to relate to a channel block of N-methyl-D-aspartate receptors. An understanding of how these compounds interact with the lipid bilayer is thus of great interest. We used molecular-dynamics simulations to calculate the potential of mean force of adamantanes in a lipid bilayer. Our results demonstrate a preference for the interfacial region of the lipid bilayer for both protonated and deprotonated species, with the protonated species proving significantly more favorable. However, the protonated species have a large free-energy barrier in the center of the membrane. In contrast, there is no barrier (compared with aqueous solution) at the center of the bilayer for deprotonated species, suggesting that the permeant species is indeed the neutral form, as commonly assumed. We discuss the results with respect to proposed mechanisms of action and implications for drug-delivery in general.     

Molecular dynamics simulation of the M2 helices within the nicotinic acetylcholine receptor transmembrane domain: structure and collective motions

Multiple nanosecond duration molecular dynamics simulations were performed on the transmembrane region of the Torpedo nicotinic acetylcholine receptor embedded within a bilayer mimetic octane slab. The M2 helices and M2-M3 loop regions were free to move, whereas the outer (M1, M3, M4) helix bundle was backbone restrained. The M2 helices largely retain their hydrogen-bonding pattern throughout the simulation, with some distortions in the helical end and loop regions. All of the M2 helices exhibit bending motions, with the hinge point in the vicinity of the central hydrophobic gate region (corresponding to residues alphaL251 and alphaV255). The bending motions of the M2 helices lead to a degree of dynamic narrowing of the pore in the region of the proposed hydrophobic gate. Calculations of Born energy profiles for various structures along the simulation trajectory suggest that the conformations of the M2 bundle sampled correspond to a closed conformation of the channel. Principal components analyses of each of the M2 helices, and of the five-helix M2 bundle, reveal concerted motions that may be relevant to channel function. Normal mode analyses using the anisotropic network model reveal collective motions similar to those identified by principal components analyses.     

Protonation of histidine and histidine-tryptophan interaction in the activation of the M2 ion channel from influenza a virus

The M2 protein of influenza A virus forms a homotetramer ion channel in the lipid membrane. The channel is specific for proton conductance and is activated by low pH with a transition midpoint at pH 5.7. We have studied the structure of the transmembrane domain of the M2 ion channel by using UV resonance Raman spectroscopy, with special attention to the side chains of histidine (His37) and tryptophan (Trp41) residues. The Raman spectra provide direct evidence that the imidazole ring of His37 is protonated upon channel activation at low pH. Concomitantly, the UV resonance Raman scattering from Trp41 shows an unusual intensity change, which is ascribed to a cation-pi interaction between the protonated (cationic) imidazole ring of His37 and the indole ring of Trp41. The protonation of His37 and the Raman intensity change of Trp41 do not occur in the presence of amantadine that blocks the M2 ion channel. These observations clearly show that the protonation of His37 and concomitant cation-pi interaction with Trp41 is a key step in the activation of the M2 ion channel. The His37-Trp41 interaction associated with the channel activation is explained by assuming a conformational transition of His37 induced by electrostatic repulsion among the protonated imidazole rings of four His37 residues in the tetramer channel. Trp41 may play a role in stabilizing the channel open state through cation-pi interaction with His37. A molecular model for the activation of M2 ion channel is proposed on the basis of the gating mechanism.     

Voltage-dependent insertion of alamethicin at phospholipid/water and octane/water interfaces

Understanding the binding and insertion of peptides in lipid bilayers is a prerequisite for understanding phenomena such as antimicrobial activity and membrane-protein folding. We describe molecular dynamics simulations of the antimicrobial peptide alamethicin in lipid/water and octane/water environments, taking into account an external electric field to mimic the membrane potential. At cis-positive potentials, alamethicin does not insert into a phospholipid bilayer in 10 ns of simulation, due to the slow dynamics of the peptide and lipids. However, in octane N-terminal insertion occurs at field strengths from 0.33 V/nm and higher, in simulations of up to 100 ns duration. Insertion of alamethicin occurs in two steps, corresponding to desolvation of the Gln7 side chain, and the backbone of Aib10 and Gly11. The proline induced helix kink angle does not change significantly during insertion. Polyalanine and alamethicin form stable helices both when inserted in octane and at the water/octane interface, where they partition in the same location. In water, both polyalanine and alamethicin partially unfold in multiple simulations. We present a detailed analysis of the insertion of alamethicin into the octane slab and the influence of the external field on the peptide structure. Our findings give new insight into the mechanism of channel formation by alamethicin and the structure and dynamics of membrane-associated helices.     

Coarse-grained molecular dynamics of tetrameric transmembrane peptide bundles within a lipid bilayer

The conformations of model transmembrane peptides are studied to understand the structural and dynamical aspects of tetrameric bundles using a series of coarse grain (CG) molecular dynamics (MD) simulations since membrane proteins play a crucial role in cell function. In this work, two different amphipathic models have been constructed using similar hydrophobic/hydrophilic characteristics with two structurally distinct morphologies to evaluate the effect of roughness and hydrophilic topology on the structure of tetrameric bundles, one class that forms an ion-channel and one class that does not. Free energy calculations of typical amphipathic peptide topologies show that using a relatively smooth surface morphology allows for a stable conformation of the tetramer bundle in a diamond formation. However, the model with side chains attached to the core in order to roughen the surface has a stable square tetramer bundle which is consistent with experimental data and all-atom (AA) MD simulations. Comparisons of the CG simulations with AA MD simulations are in reasonable agreement with the formation of tetrameric homo-oligomers, partitioning within the lipid bilayer and tilt angle with respect to the bilayer normal. We concluded that a square or diamond shape tetrameric homo-oligomers could be stabilized by rational design of the peptide morphology and topology of the surface, thus allowing us to tune the permeability of the bundle or channel.     

Understanding pH-dependent selectivity of alamethicin K18 channels by computer simulation

Alamethicin K18 is a covalently linked alamethicin dimer in which the glutamine residue at position 18 in each helix has been replaced by a lysine residue. As described in previous work, channels formed by this peptide show pH-dependent selectivity. The maximum anion selectivity of the putative octameric conducting state is obtained at pH 7 or lower. Inasmuch as no change in selectivity is seen between pH 7 and pH 3, and because protons are expected to be in equilibrium with the open state of the channel during a selectivity measurement, the channel is believed to be fully charged (i.e., all eight lysines protonated) at pH 7. In an effort to understand how such a highly charged channel structure is stable in membranes and why it is not more selective for anions, we have performed a number of computer simulations of the system. Molecular dynamics simulations of 10 ns each of the octameric bundle in a lipid bilayer environment are presented, with either zero, four, or eight lysines charged in the absence of salt, and with eight lysines charged in the presence of 0.5 M and 1 M KCl. When no salt is present and all lysines are charged, on average 1.9 Cl(-) ions are inside the channel and the channel significantly deforms. With 0.5 M KCl present, 2.9 Cl(-) ions are inside the channel. With 1 M KCl present, four Cl(-) ions are present and the channel maintains a regular structure. Poisson-Boltzmann calculations on models of the octameric channel also predict an average of 2-4 Cl(-) ions near the lysine residues as a function of ionic strength. These counterions lower the apparent charge of the channel, which may underlie the decrease in selectivity observed experimentally with increasing salt concentrations. We suggest that to increase the selectivity of Alm K18 channels, positive charges could be engineered in a narrower part of the channel.     

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.     

Molecular dynamics study of the M412 intermediate of bacteriorhodopsin.

Molecular dynamics simulations have been carried out to study the M412 intermediate of bacteriorhodopsin's (bR) photocycle. The simulations start from two simulated structures for the L550 intermediate of the photocycle, one involving a 13-cis retinal with strong torsions, the other a 13,14-dicis retinal, from which the M412 intermediate is initiated through proton transfer to Asp-85. The simulations are based on a refined structure of bR568 obtained through all-atom molecular dynamics simulations and placement of 16 waters inside the protein. The structures of the L550 intermediates were obtained through simulated photoisomerization and subsequent molecular dynamics, and simulated annealing. Our simulations reveal that the M412 intermediate actually comprises a series of conformations involving 1) a motion of retinal; 2) protein conformational changes; and 3) diffusion and reconfiguration of water in the space between the retinal Schiff base nitrogen and the Asp-96 side group. (1) turns the retinal Schiff base nitrogen from an early orientation toward Asp-85 to a late orientation toward Asp-96; (2) disconnects the hydrogen bond network between retinal and Asp-85 and tilts the helix F of bR, enlarging bR's cytoplasmic channel; (3) adds two water molecules to the three water molecules existing in the cytoplasmic channel at the bR568 stage and forms a proton conduction pathway. The conformational change (2) of the protein involves a 60 degrees bent of the cytoplasmic side of helix F and is induced through a break of a hydrogen bond between Tyr-185 and a water-side group complex in the counterion region.     

Assembly of Nsp1 Nucleoporins Provides Insight into Nuclear Pore Complex Gating

Nuclear pore complexes (NPCs) form gateways for material transfer across the nuclear envelope of eukaryotic cells. Disordered proteins, rich in phenylalanine-glycine repeat motifs (FG-nups), form the central transport channel. Understanding how nups are arranged in the interior of the NPC may explain how NPC functions as a selectivity filter for transport of large molecules and a sieve-like filter for diffusion of small molecules (< or ). We employed molecular dynamics to model the structures formed by various assemblies of one kind of nup, namely the 609-aa-long FG domain of Nsp1 (Nsp1-FG). The simulations started from different initial conformations and geometrical arrangements of Nsp1-FGs. In all cases Nsp1-FGs collectively formed brush-like structures with bristles made of bundles of 2–27 nups, however, the bundles being cross-linked through single nups leaving one bundle and joining a nearby one. The degree of cross-linking varies with different initial nup conformations and arrangements. Structural analysis reveals that FG-repeats of the nups not only involve formation of bundle structures, but are abundantly present in cross-linking regions where the epitopes of FG-repeats are highly accessible. Large molecules that are assisted by transport factors (TFs) are selectively transported through NPC apparently by binding to FG-nups through populated FG-binding pockets on the TF surface. Therefore, our finding suggests that TFs bind concertedly to multiple FGs in cross-linking regions and break-up the bundles to create wide pores for themselves and their cargoes to pass. In addition, the cross-linking between Nsp1-FG bundles, arising from simulations, is found to set a molecular size limit of < for passive diffusion of molecules. Our simulations suggest that the NPC central channel, near the periphery where tethering of nups is dominant, features brush-like moderately cross-linked bundles, but in the central region, where tethering loses its effect, features a sieve-like structure of bundles and frequent cross-links.     

Activation and Proton Transport Mechanism in Influenza A M2 Channel

Molecular dynamics trajectories 2 μs in length have been generated for the pH-activated, tetrameric M2 proton channel of the influenza A virus in all protonation states of the pH sensor located at the His³⁷ tetrad. All simulated structures are in very good agreement with high-resolution structures. Changes in the channel caused by progressive protonation of His³⁷ provide insight into the mechanism of proton transport. The channel is closed at both His³⁷ and Trp⁴¹ sites in the singly and doubly protonated states, but it opens at Trp⁴¹ upon further protonation. Anions access the charged His³⁷ and by doing so stabilize the protonated states of the channel. The narrow opening at the His³⁷ site, further blocked by anions, is inconsistent with the water-wire mechanism of proton transport. Instead, conformational interconversions of His³⁷ correlated with hydrogen bonding to water molecules indicate that these residues shuttle protons in high-protonation states. Hydrogen bonds between charged and uncharged histidines are rare. The valve at Val²⁷ remains on average quite narrow in all protonation states but fluctuates sufficiently to support water and proton transport. A proton transport mechanism in which the channel, depending on pH, opens at either the histidine or valine gate is only partially supported by the simulations.     

KcsA closed and open: modelling and simulation studies

Bacterial homologues of mammalian potassium channels provide structures of two states of a gated K channel. Thus, the crystal structure of KcsA represents a closed state whilst that of MthK represents an open state. Using homology modelling and molecular dynamics simulations we have built a model of the transmembrane domain of KcsA in an open state and have compared its conformational stability with that of the same domain of KcsA in a closed state. Approximate Born energy calculations of monovalent cations within the two KcsA channel states suggest that the intracellular hydrophobic gate in the closed state provides a barrier of height ~5 kT to ion permeation, whilst in the open state the barrier is absent. Simulations (10 ns duration) in an octane slab (a simple membrane mimetic) suggest that closed- and open-state models are of comparable conformational stability, both exhibiting conformational drifts of ~3.3 Å C RMSD relative to the respective starting models. Substantial conformational fluctuations are observed in the intracellular gate region during both simulations (closed state and open state). In the simulation of open-state KcsA, rapid (<5 ns) exit of all three K⁺ ions occurs through the intracellular mouth of the channel. Helix kink and swivel motion is observed at the molecular hinge formed by residue G99 of the M2 helix. This motion is more substantial for the open- than for the closed-state model of the channel.     

Activation and Proton Transport Mechanism in Influenza A M2 Channel

Molecular dynamics trajectories 2 μs in length have been generated for the pH-activated, tetrameric M2 proton channel of the influenza A virus in all protonation states of the pH sensor located at the His³⁷ tetrad. All simulated structures are in very good agreement with high-resolution structures. Changes in the channel caused by progressive protonation of His³⁷ provide insight into the mechanism of proton transport. The channel is closed at both His³⁷ and Trp⁴¹ sites in the singly and doubly protonated states, but it opens at Trp⁴¹ upon further protonation. Anions access the charged His³⁷ and by doing so stabilize the protonated states of the channel. The narrow opening at the His³⁷ site, further blocked by anions, is inconsistent with the water-wire mechanism of proton transport. Instead, conformational interconversions of His³⁷ correlated with hydrogen bonding to water molecules indicate that these residues shuttle protons in high-protonation states. Hydrogen bonds between charged and uncharged histidines are rare. The valve at Val²⁷ remains on average quite narrow in all protonation states but fluctuates sufficiently to support water and proton transport. A proton transport mechanism in which the channel, depending on pH, opens at either the histidine or valine gate is only partially supported by the simulations.     

Molecular Dynamics Simulations of Homo-oligomeric Bundles Embedded Within a Lipid Bilayer

Using molecular dynamics simulations, we studied the structure, interhelix interactions, and dynamics of transmembrane proteins. Specifically, we investigated homooligomeric helical bundle systems consisting of synthetic α-helices with either the sequence Ac-(LSLLLSL)₃-NH₂ (LS2) or Ac-(LSSLLSL)₃-NH₂ (LS3). The LS2 and LS3 helical peptides are designed to have amphipathic characteristics that form ion channels in membrane. We simulated bundles containing one to six peptides that were embedded in palmitoyl-oleoyl-phosphatidylcholine (POPC) lipid bilayer and placed between two lamellae of water. We aim to provide a fundamental understanding of how amphipathic helical peptides interact with each other and their dynamical behaviors in different homooligomeric states. To understand structural properties, we examined the helix lengths, tilt angles of individual helices and the entire bundle, interhelix distances, interhelix cross-angles, helix hydrophobic-to-hydrophilic vector projections, and the average number of interhelix hydrophilic (serine–serine) contacts lining the pore of the transmembrane channel. To analyze dynamical properties, we calculated the rotational autocorrelation function of each helix and the cross-correlation of the rotational velocity between adjacent helices. The observed structural and dynamical characteristics show that higher order bundles containing four to six peptides are composed of multiple lower order bundles of one to three peptides. For example, the LS2 channel was found to be stable in a tetrameric bundle composed of a “dimer of dimers.” In addition, we observed that there is a minimum of two strong hydrophilic contacts between a pair of adjacent helices in the dimer to tetramer systems and only one strong hydrophilic interhelix contact in helix pairs of the pentamer and hexamer systems. We believe these results are general and can be applied to more complex ion channels, providing insight into ion channel stability and assembly.     

Molecular Dynamics Simulations of Homo-oligomeric Bundles Embedded Within a Lipid Bilayer

Using molecular dynamics simulations, we studied the structure, interhelix interactions, and dynamics of transmembrane proteins. Specifically, we investigated homooligomeric helical bundle systems consisting of synthetic α-helices with either the sequence Ac-(LSLLLSL)₃-NH₂ (LS2) or Ac-(LSSLLSL)₃-NH₂ (LS3). The LS2 and LS3 helical peptides are designed to have amphipathic characteristics that form ion channels in membrane. We simulated bundles containing one to six peptides that were embedded in palmitoyl-oleoyl-phosphatidylcholine (POPC) lipid bilayer and placed between two lamellae of water. We aim to provide a fundamental understanding of how amphipathic helical peptides interact with each other and their dynamical behaviors in different homooligomeric states. To understand structural properties, we examined the helix lengths, tilt angles of individual helices and the entire bundle, interhelix distances, interhelix cross-angles, helix hydrophobic-to-hydrophilic vector projections, and the average number of interhelix hydrophilic (serine–serine) contacts lining the pore of the transmembrane channel. To analyze dynamical properties, we calculated the rotational autocorrelation function of each helix and the cross-correlation of the rotational velocity between adjacent helices. The observed structural and dynamical characteristics show that higher order bundles containing four to six peptides are composed of multiple lower order bundles of one to three peptides. For example, the LS2 channel was found to be stable in a tetrameric bundle composed of a “dimer of dimers.” In addition, we observed that there is a minimum of two strong hydrophilic contacts between a pair of adjacent helices in the dimer to tetramer systems and only one strong hydrophilic interhelix contact in helix pairs of the pentamer and hexamer systems. We believe these results are general and can be applied to more complex ion channels, providing insight into ion channel stability and assembly.     

Viral ion channels: molecular modeling and simulation

In a number of membrane-bound viruses, ion channels are formed by integral membrane proteins. These channel proteins include M2 from influenza A, NB from influenza B, and, possibly, Vpu from HIV-1. M2 is important in facilitating uncoating of the influenza A viral genome and is the target of amantadine, an anti-influenza drug. The biological roles of NB and Vpu are less certain. In all cases, the protein contains a single transmembrane alpha-helix close to its N-terminus. Channels can be formed by homo-oligomerization of these proteins, yielding bundles of transmembrane helices that span the membrane and surround a central ion-permeable pore. Molecular modeling may be used to integrate and interpret available experimental data concerning the structure of such transmembrane pores. This has proved successful for the M2 channel domain, where two independently derived models are in agreement with one another, and with solid-state nuclear magnetic resonance (NMR) data. Simulations based on channel models may yield insights into possible ion conduction and selectivity mechanisms.