Turgut Baştuğ

Gramicidin A channel as a test ground for molecular dynamics force fields

We use the well-known structural and functional properties of the gramicidin A channel to test the appropriateness of force fields commonly used in molecular dynamics (MD) simulations of ion channels. For this purpose, the high-resolution structure of the gramicidin A dimer is embedded in a dimyristoylphosphatidylcholine bilayer, and the potential of mean force of a K(+) ion is calculated along the channel axis using the umbrella sampling method. Calculations are performed using two of the most common force fields in MD simulations: CHARMM and GROMACS. Both force fields lead to large central barriers for K(+) ion permeation, that are substantially higher than those deduced from the physiological data by inverse methods. In long MD simulations lasting over 60 ns, several ions are observed to enter the binding site but none of them crossed the channel despite the presence of a large driving field. The present results, taken together with many earlier studies, highlights the shortcomings of the standard force fields used in MD simulations of ion channels and calls for construction of more appropriate force fields for this purpose.

Potential of mean force calculations of ligand binding to ion channels from Jarzynski's equality and umbrella sampling.

Potential of mean force (PMF) calculations provide a reliable method for determination of the absolute binding free energies for protein-ligand systems. The common method used for this purpose -- umbrella sampling with weighted histogram analysis -- is computationally very laborious, which limits its applications. Recently, a much simpler alternative for PMF calculations has become available, namely, using Jarzynskis equality in steered molecular dynamics simulations. So far, there have been a few comparisons of the two methods and mostly in simple systems that do not reflect the complexities of protein-ligand systems. Here, we use both methods to calculate the PMF for ion permeation and ligand binding to ion channels. Comparison of results indicate that Jarzynskis method suffers from relaxation problems in complex systems and would require much longer simulation times to yield reliable PMFs for protein-ligand systems.

Position of the Third Na+ Site in the Aspartate Transporter GltPh and the Human Glutamate Transporter, EAAT1

Glutamate transport via the human excitatory amino acid transporters is coupled to the co-transport of three Na+ ions, one H+ and the counter-transport of one K+ ion. Transport by an archaeal homologue of the human glutamate transporters, GltPh, whose three dimensional structure is known is also coupled to three Na+ ions but only two Na+ ion binding sites have been observed in the crystal structure of GltPh. In order to fully utilize the GltPh structure in functional studies of the human glutamate transporters, it is essential to understand the transport mechanism of GltPh and accurately determine the number and location of Na+ ions coupled to transport. Several sites have been proposed for the binding of a third Na+ ion from electrostatic calculations and molecular dynamics simulations. In this study, we have performed detailed free energy simulations for GltPh and reveal a new site for the third Na+ ion involving the side chains of Threonine 92, Serine 93, Asparagine 310, Aspartate 312, and the backbone of Tyrosine 89. We have also studied the transport properties of alanine mutants of the coordinating residues Threonine 92 and Serine 93 in GltPh, and the corresponding residues in a human glutamate transporter, EAAT1. The mutant transporters have reduced affinity for Na+ compared to their wild type counterparts. These results confirm that Threonine 92 and Serine 93 are involved in the coordination of the third Na+ ion in GltPh and EAAT1.

The electronic structure and properties of group 8 oxides MO4, where M=Ru, Os, and Element 108, Hs

Fully relativistic density functional calculations have been performed for group 8 tetroxides MO4, where M=Ru, Os, and element 108, Hs. The electronic structure analysis has shown HsO4 to be very similar to OsO4, with the covalence and stability increasing from OsO4 to HsO4. Using models of atom-slab interactions, adsorption enthalpies of RuO4 and HsO4 on the quartz surface have been calculated using some models of physisorption. The volatility of the single species was shown to have the following trend, RuO4<OsO4⩽HsO4, with differences in the adsorption enthalpies between the species being almost within the experimental uncertainty of ±1.5 kJ/mol.

Intermetallic compounds of the heaviest elements: the electronic structure and bonding of dimers of element 112 and its homolog Hg

Abstract Fully relativistic (four-component) density-functional calculations were performed for the element 112 dimers (112)X (X = Pd, Cu, Ag and Au) and those of its lighter homolog, Hg. A relatively small decrease of about 15–20 kJ/mol in bonding was found from the HgX to (112)X compounds. Respectively, the bond lengths were increased by 0.06 A on the average. The Mulliken population analysis has shown this effect to be a result of a decreasing contribution of the relativistically stabilized 7s-AO of element 112 to bonding. The following trend in the binding energies was predicted for (112)X as a function of X: Pd >Cu>Au>Ag, exactly as the trend obtained experimentally for adsorption of Hg on the corresponding metal surfaces.

Free energy simulations of single and double ion occupancy in gramicidin A

Simultaneous occupancy of the two binding sites in gramicidin A by monovalent cations is a well known property of this channel, but the energetic feasibility of this process in molecular dynamics simulations has not been established so far. Here the authors study the energetics of single and double ion occupancy in gramicidin A by constructing the potential of mean force for single and pair of cations. As representatives of small and large ions, they consider both Na+ and K+ ions in the calculations. Binding constants of ions are estimated from the free energy profiles. Comparisons with the experimental results indicate 3-4 kT discrepancy in the binding energies. They also study the coordination of the ions in their respective binding sites and the dynamic behavior of the channel water during the double ion binding process.

Role of the dielectric constants of membrane proteins and channel water in ion permeation.

Using both analytical solutions obtained from simplified systems and numerical results from more realistic cases, we investigate the role played by the dielectric constant of membrane proteins epsilon(p) and pore water epsilon(w) in permeation of ions across channels. We show that the boundary and its curvature are the crucial factors in determining how an ions potential energy depends on the dielectric constants near an interface. The potential energy of an ion outside a globular protein has a dominant 1/epsilon(w) dependence, but this becomes 1/epsilon(p) for an ion inside a cavity. For channels, where the boundaries are in between these two extremes, the situation is more complex. In general, we find that variations in epsilon(w) have a much larger impact on the potential energy of an ion compared to those in epsilon(p). Therefore a better understanding of the effective epsilon(w) values employed in channel models is desirable. Although the precise value of epsilon(p) is not a crucial determinant of ion permeation properties, it still needs to be chosen carefully when quantitative comparisons with data are made.

Free Energy Simulations of Ligand Binding to the Aspartate Transporter GltPh

Glutamate/Aspartate transporters cotransport three Na(+) and one H(+) ions with the substrate and countertransport one K(+) ion. The binding sites for the substrate and two Na(+) ions have been observed in the crystal structure of the archeal homolog Glt(Ph), while the binding site for the third Na(+) ion has been proposed from computational studies and confirmed by experiments. Here we perform detailed free energy simulations of Glt(Ph), giving a comprehensive characterization of the substrate and ion binding sites, and calculating their binding free energies in various configurations. Our results show unequivocally that the substrate binds after the binding of two Na(+) ions. They also shed light into Asp/Glu selectivity of Glt(Ph), which is not observed in eukaryotic glutamate transporters.

Comparative Study of the Energetics of Ion Permeation in Kv1.2 and KcsA Potassium Channels

Biological ion channels rely on a multi-ion transport mechanism for fast yet selective permeation of ions. The crystal structure of the KcsA potassium channel provided the first microscopic picture of this process. A similar mechanism is assumed to operate in all potassium channels, but the validity of this assumption has not been well investigated. Here, we examine the energetics of ion permeation in Shaker Kv1.2 and KcsA channels, which exemplify the six-transmembrane voltage-gated and two-transmembrane inward-rectifier channels. We study the feasibility of binding a third ion to the filter and the concerted motion of ions in the channel by constructing the potential of mean force for K(+) ions in various configurations. For both channels, we find that a pair of K(+) ions can move almost freely within the filter, but a relatively large free-energy barrier hinders the K(+) ion from stepping outside the filter. We discuss the effect of the CMAP dihedral energy correction that was recently incorporated into the CHARMM force field on ion permeation dynamics.

Importance of the Peptide Backbone Description in Modeling the Selectivity Filter in Potassium Channels

A dihedral energy correction (CMAP) term has been recently included in the CHARMM force field to obtain a more accurate description of the peptide backbone. Its importance in improving dynamical properties of proteins and preserving their stability in long molecular-dynamics simulations has been established for several globular proteins. Here we investigate its role in maintaining the structure and function of two potassium channels, Shaker K(v)1.2 and KcsA, by performing molecular-dynamics simulations with and without the CMAP correction in otherwise identical systems. We show that without CMAP, it is not possible to maintain the experimentally observed orientations of the carbonyl groups in the selectivity filter in Shaker, and the channel loses its selectivity property. In the case of KcsA, the channel retains some selectivity even without CMAP because the carbonyl orientations are relatively better preserved compared to Shaker.