Frank R. Beierlein

A Simple QM/MM Approach for Capturing Polarization Effects in Protein-Ligand Binding Free Energy Calculations

We present a molecular simulation protocol to compute free energies of binding, which combines a QM/MM correction term with rigorous classical free energy techniques, thereby accounting for electronic polarization effects. Relative free energies of binding are first computed using classical force fields, Monte Carlo sampling, and replica exchange thermodynamic integration. Snapshots of the configurations at the end points of the perturbation are then subjected to DFT-QM/MM single-point calculations using the B3LYP functional and a range of basis sets. The resulting quantum mechanical energies are then processed using the Zwanzig equation to give free energies incorporating electronic polarization. Our approach is conceptually simple and does not require tightly coupled QM and MM software. The method has been validated by calculating the relative free energies of hydration of methane and water and the relative free energy of binding of two inhibitors of cyclooxygenase-2. Closed thermodynamic cycles are obtained across different pathways, demonstrating the correctness of the technique, although significantly more sampling is required for the protein-ligand system. Our method offers a simple and effective way to incorporate quantum mechanical effects into computed free energies of binding.

Quantum mechanical/molecular mechanical (QM/MM) docking: an evaluation for known test systems

A combined quantum mechanical/molecular mechanical (QM/MM) docking approach for the investigation of protein-inhibitor complexes is presented. Starting points for QM/MM optimizations are generated with AutoDock. The subsequent semiempirical AMI QM/MM optimization of the complex obtained by the docking procedure gives a more detailed description of the binding mode and the electronic properties of the ligand. As we use a flexible protein environment in the QM/MM optimizations, we are able to simulate limited structural changes of the enzyme upon binding a ligand, even within a simple geometry optimization. The method was validated using a set of structurally known protein-inhibitor complexes, whose crystallographic data were taken from the Protein Data Bank. In addition to protein structures taken directly from complexes with the inhibitors, structures of uncomplexed HIV-1-protease and thrombin were also used successfully for QM/MM docking experiments. By comparing the resulting structures with those obtained using protein structures from protein-inhibitor complexes, we find that the method is able to simulate the effect of the induced fit when a simple optimization is adequate to reproduce the protein movement. Describing the ligand quantum mechanically gives a detailed view of its electronic properties, for example its polarization within the active site of the enzyme. This study suggests strongly that a QM/MM molecular dynamics approach will be able to simulate the induced fit in general cases.

Predicting the Effects of Basepair Mutations in DNA-Protein Complexes by Thermodynamic Integration

Thermodynamically rigorous free energy methods in principle allow the exact computation of binding free energies in biological systems. Here, we use thermodynamic integration together with molecular dynamics simulations of a DNA-protein complex to compute relative binding free energies of a series of mutants of a protein-binding DNA operator sequence. A guanine-cytosine basepair that interacts strongly with the DNA-binding protein is mutated into adenine-thymine, cytosine-guanine, and thymine-adenine. It is shown that basepair mutations can be performed using a conservative protocol that gives error estimates of ∼10% of the change in free energy of binding. Despite the high CPU-time requirements, this work opens the exciting opportunity of being able to perform basepair scans to investigate protein-DNA binding specificity in great detail computationally.

Molecular Dynamics Simulations of Liquid Phase Interfaces: Understanding the Structure of the Glycerol/Water−Dodecane System

Modern spectroscopic techniques such as time-resolved second-harmonic-generation spectroscopy allow molecules to be examined selectively directly at phase interfaces. Two-phase systems formed by glycerol/water and alkane layers have previously been studied by time-resolved second-harmonic-generation spectroscopic measurements. In this molecular dynamics study, a triphenylmethane dye was inserted at the glycerol/water-alkane interface and was used as a probe for local properties such as viscosity. We now show how extensive simulations over a wide range of concentrations can be used to obtain a detailed view of the molecular structure at the glycerol/water-alkane interface. Glycerol is accumulated in a double layer adjacent to the alkane interface, which results in increased viscosity of the glycerol/water phase in the direct vicinity of the interface. We also show that conformational ensembles created by classical molecular-dynamics simulations can serve as input for QM/MM calculations, yielding further information such as transition dipoles, which can be compared with spectroscopic measurements.

Carboxylate Ion Pairing with Alkali-Metal Ions for β-Lactoglobulin and Its Role on Aggregation and Interfacial Adsorption

We report a combined experimental and computational study of the whey protein β-lactoglobulin (BLG) in different electrolyte solutions. Vibrational sum-frequency generation (SFG) and ellipsometry were used to investigate the molecular structure of BLG modified air-water interfaces as a function of LiCl, NaCl, and KCl concentrations. Molecular dynamics (MD) simulations and thermodynamic integration provided details of the ion pairing of protein surface residues with alkali-metal cations. Our results at pH 6.2 indicate that BLG at the air-water interface forms mono- and bilayers preferably at low and high ionic strength, respectively. Results from SFG spectroscopy and ellipsometry are consistent with intimate ion pairing of alkali-metal cations with aspartate and glutamate carboxylates, which is shown to be more effective for smaller cations (Li(+) and Na(+)). MD simulations show not only carboxylate-alkali-metal ion pairs but also ion multiplets with the alkali-metal ion in a bridging position between two or more carboxylates. Consequently, alkali-metal cations can bridge carboxylates not only within a monomer but also between monomers, thus providing an important dimerization mechanism between hydrophilic surface patches.

A test of improved force field parameters for urea: molecular-dynamics simulations of urea crystals

Molecular-dynamics (MD) simulations of urea crystals of different shapes (cubic, rectangular prismatic, and sheet) have been performed using our previously published force field for urea. This force field has been validated by calculating values for the cohesive energy, sublimation temperature, and melting point from the MD data. The cohesive energies computed from simulations of cubic and rectangular prismatic urea crystals in vacuo at 300 K agreed very well with the experimental sublimation enthalpies reported at 298 K. We also found very good agreement between the melting points as observed experimentally and from simulations. Annealing the crystals just below the melting point leads to reconstruction to form crystal faces that are consistent with experimental observations. The simulations reveal a melting mechanism that involves surface (corner/edge) melting well below the melting point, and rotational disordering of the urea molecules in the corner/edge regions of the crystal, which then facilitates the translational motion of these molecules.

Computer Simulations of Enzyme Reaction Mechanisms: Simulation of Protein Spectra

Enzymes play a key role in modern pharmaceutical research because they represent targets for the design of new drugs. In addition to the classical approach of inhibiting an enzyme by blocking its binding site with an inhibitor, the level of gene expression is now moving into the focus of interest. An important system for the investigation of mechanisms of transcriptional control is the Tet repressor/tet operator (TetR/tetO) system. We employ a combined classical/quantum mechanical approach to model the structure and the spectroscopic properties of the TetR-tetracycline complex. As our methods are based on semiempirical molecular orbital theory, we have also developed a parallel pseudodiagonalization technique for the major computational step in such calculations. The parallel pseudodiagonalizer gives acceptable performance for up to about eight processors.

DNA-Dye-Conjugates: Conformations and Spectra of Fluorescence Probes

Extensive molecular-dynamics (MD) simulations have been used to investigate DNA-dye and DNA-photosensitizer conjugates, which act as reactants in templated reactions leading to the generation of fluorescent products in the presence of specific desoxyribonucleic acid sequences (targets). Such reactions are potentially suitable for detecting target nucleic acids in live cells by fluorescence microscopy or flow cytometry. The simulations show how the attached dyes/photosensitizers influence DNA structure and reveal the relative orientations of the chromophores with respect to each other. Our results will help to optimize the reactants for the templated reactions, especially length and structure of the spacers used to link reporter dyes or photosensitizers to the oligonucleotides responsible for target recognition. Furthermore, we demonstrate that the structural ensembles obtained from the simulations can be used to calculate steady-state UV-vis absorption and emission spectra. We also show how important quantities describing the quenching of the reporter dye via fluorescence resonance energy transfer (FRET) can be calculated from the simulation data, and we compare these for different relative chromophore geometries.