Stefan M. Kast

Potential energy function for apatites

An empirical potential energy function for fluor- and for hydroxyapatite is formulated and parametrised. The parameter optimisation involves a hierarchy of reference data and techniques comprising of quantum-chemical calculations for Coulomb interactions and intramolecular contributions, as well as experimental data and molecular dynamics simulations for the remaining nonbonded parameters. For fluorapatite both a flexible and a rigid phosphate model are derived, while for hydroxyapatite only the rigid variant is determined. Simulations with the final models reproduce the experimental crystal parameters within less than 1% deviation for a wide range of temperatures between 73 and 1273 K. In the case of flexible fluorapatite the computed and the experimental infrared spectra at 300 K agree excellently.

Closed-form expressions of the chemical potential for integral equation closures with certain bridge functions

A general, path-independent expression is derived for the excess chemical potential of integral equation closure approximations that contain a bridge function which depends on a renormalized indirect correlation function. Closed-form results are obtained for various cases, among them a partial series expansion of the hypernetted chain closure.

Constant temperature molecular dynamics simulations by means of a stochastic collision model. I. Noninteracting particles

A new stochastic collision model for molecular dynamics (MD) simulations at constant temperature is presented. It is based on impulsive collisions between system particles and heat bath particles of finite masses. With the algorithm one can switch between Langevin‐type and Andersen‐type dynamics by changing only one control parameter. The method is implemented in the simple Verlet scheme for the numerical calculation of the Newtonian equations of motion. The case of an ensemble of noninteracting particles subjected to the heat bath is considered. Analytical expressions for stationary probability densities, autocorrelation functions, and diffusion coefficients are derived. The predicitions agree excellently with the results of MD simulations.

Quantum Chemistry in Solution by Combining 3D Integral Equation Theory with a Cluster Embedding Approach

Free energy changes associated with chemical reactions in solution are treated by integral equation theory in the form of the 3D reference interaction site model (RISM) in combination with quantum-chemical calculations via an embedded cluster approach (EC-RISM). The electronic structure of the solute is computed self-consistently with the solvent structure by mapping the charge distribution of the solvent onto a set of discrete background point charges that are added to the molecular Hamiltonian. The EC-RISM procedure yields chemical accuracy in free energy predictions for several benchmark systems without adjusting empirical parameters. We apply the method to the standard reaction free energy for the gauche-trans equilibrium of 1,2-dichloroethane in water and to pKa shift calculations for trifluoroacetic acid/acetic acid and 4-nitroaniline/aniline in water.

The proapoptotic influenza A virus protein PB1-F2 forms a nonselective ion channel

Background PB1-F2 is a proapoptotic influenza A virus protein of approximately 90 amino acids in length that is located in the nucleus, cytosol and in the mitochondria membrane of infected cells. Previous studies indicated that the molecule destabilizes planar lipid bilayers and has a strong inherent tendency for multimerization. This may be correlate with its capacity to induce mitochondrial membrane depolarization. Methodology/Principal Findings Here, we investigated whether PB1-F2 is able to form ion channels within planar lipid bilayers and microsomes. For that purpose, a set of biologically active synthetic versions of PB1-F2 (sPB1-F2) derived from the IAV isolates A/Puerto Rico/8/34(H1N1) (IAVPR8), from A/Brevig Mission/1/1918(H1N1) (IAVSF2) or the H5N1 consensus sequence (IAVBF2) were used. Electrical and fluorimetric measurements show that all three peptides generate in planar lipid bilayers or in liposomes, respectively, a barely selective conductance that is associated with stochastic channel type fluctuations between a closed state and at least two defined open states. Unitary channel fluctuations were also generated when a truncated protein comprising only the 37 c-terminal amino acids of sPB1-F2 was reconstituted in bilayers. Experiments were complemented by extensive molecular dynamics simulations of the truncated fragment in a lipid bilayer. The results indicate that the c-terminal region exhibits a slightly bent helical fold, which is stable and remains embedded in the bilayer for over 180 ns. Conclusion/Significance The data support the idea that PB1-F2 is able to form protein channel pores with no appreciable selectivity in membranes and that the c-terminus is important for this function. This information could be important for drug development.

Targeting Drug Resistance in EGFR with Covalent Inhibitors: A Structure-Based Design Approach.

Receptor tyrosine kinases represent one of the prime targets in cancer therapy, as the dysregulation of these elementary transducers of extracellular signals, like the epidermal growth factor receptor (EGFR), contributes to the onset of cancer, such as non-small cell lung cancer (NSCLC). Strong efforts were directed to the development of irreversible inhibitors and led to compound CO-1686, which takes advantage of increased residence time at EGFR by alkylating Cys797 and thereby preventing toxic effects. Here, we present a structure-based approach, rationalized by subsequent computational analysis of conformational ligand ensembles in solution, to design novel and irreversible EGFR inhibitors based on a screening hit that was identified in a phenotype screen of 80 NSCLC cell lines against approximately 1500 compounds. Using protein X-ray crystallography, we deciphered the binding mode in engineered cSrc (T338M/S345C), a validated model system for EGFR-T790M, which constituted the basis for further rational design approaches. Chemical synthesis led to further compound collections that revealed increased biochemical potency and, in part, selectivity toward mutated (L858R and L858R/T790M) vs nonmutated EGFR. Further cell-based and kinetic studies were performed to substantiate our initial findings. Utilizing proteolytic digestion and nano-LC-MS/MS analysis, we confirmed the alkylation of Cys797.

Chlorella virus ATCV-1 encodes a functional potassium channel of 82 amino acids.

Chlorella virus PBCV-1 (Paramecium bursaria chlorella virus-1) encodes the smallest protein (94 amino acids, named Kcv) previously known to form a functional K+ channel in heterologous systems. In this paper, we characterize another chlorella virus encoded K+ channel protein (82 amino acids, named ATCV-1 Kcv) that forms a functional channel in Xenopus oocytes and rescues Saccharomyces cerevisiae mutants that lack endogenous K+ uptake systems. Compared with the larger PBCV-1 Kcv, ATCV-1 Kcv lacks a cytoplasmic N-terminus and has a reduced number of charged amino acids in its turret domain. Despite these deficiencies, ATCV-1 Kcv accomplishes all the major features of K+ channels: it assembles into a tetramer, is K+ selective and is inhibited by the canonical K+ channel blockers, barium and caesium. Single channel analyses reveal a stochastic gating behaviour and a voltage-dependent conductance that resembles the macroscopic I/V relationship. One difference between PBCV-1 and ATCV-1 Kcv is that the latter is more permeable to K+ than Rb+. This difference is partially explained by the presence of a tyrosine residue in the selective filter of ATCV-1 Kcv, whereas PBCV-1 Kcv has a phenylalanine. Hence, ATCV-1 Kcv is the smallest protein to form a K+ channel and it will serve as a model for studying structure-function correlations inside the potassium channel pore.

Constant temperature molecular dynamics simulations by means of a stochastic collision model. II. The harmonic oscillator

Our recently introduced stochastic method for molecular dynamics simulations at constant temperature [J. Chem. Phys. 100, 566 (1994)] which is based on impulsive collisions between system particles and heat bath particles of finite masses, is extended and analyzed for the case of an ensemble of harmonic oscillators as a simple but theoretically solvable model for interacting systems. This model case can be considered, e.g., as a single normal mode of a polyatomic molecule. Both position space properties and velocity space properties are investigated. Analytical expressions for stationary probability densities and autocorrelation functions are derived. The effect of the truncation of the Taylor series which is the basis of Verlet‐type algorithms, on the resulting temperature in position space and velocity space is quantitatively discussed. It is demonstrated that the system temperature in position space and in velocity space is controlled by complex parameter functions.

Model development for the viral Kcv potassium channel.

A computational model for the open state of the short viral Kcv potassium channel was created and tested based on homology modeling and extensive molecular-dynamics simulation in a membrane environment. Particular attention was paid to the structure of the highly flexible N-terminal region and to the protonation state of membrane-exposed lysine residues. Data from various experimental sources, NMR spectroscopy, and electrophysiology, as well as results from three-dimensional reference interaction site model integral equation theory were taken into account to select the most reasonable model among possible variants. The final model exhibits spontaneous ion transitions across the complete pore, with and without application of an external field. The nonequilibrium transport events could be induced reproducibly without abnormally large driving potential and without the need to place ions artificially at certain key positions along the transition path. The transport mechanism through the filter region corresponds to the classic view of single-file motion, which in our case is coupled to frequent exchange of ions between the innermost filter position and the cavity.

Long Distance Interactions within the Potassium Channel Pore Are Revealed by Molecular Diversity of Viral Proteins

Kcv is a 94-amino acid protein encoded by chlorella virus PBCV-1 that corresponds to the pore module of K+ channels. Therefore, Kcv can be a model for studying the protein design of K+ channel pores. We analyzed the molecular diversity generated by ∼1 billion years of evolution on kcv genes isolated from 40 additional chlorella viruses. Because the channel is apparently required for virus replication, the Kcv variants are all functional and contain multiple and dispersed substitutions that represent a repertoire of allowed sets of amino acid substitutions (from 4 to 12 amino acids). Correlations between amino acid substitutions and the new properties displayed by these channels guided site-directed mutations that revealed synergistic amino acid interactions within the protein as well as previously unknown interactions between distant channel domains. The effects of these multiple changes were not predictable from a priori structural knowledge of the channel pore.