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Insights from the First Principles Based Large Scale Protein Thermostability Calculations
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Conformational Transitions upon Ligand Binding: Holo-Structure Prediction from Apo Conformations
No full textAtomic contacts in protein structures. A detailed analysis of atomic radii, packing, and overlaps.
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Protein thermostability calculations using alchemical free energy simulations.
Full text linkAbstractThermal stability of proteins is crucial for both biotechnological and therapeutic applications. Rational protein engineering therefore frequently aims at increasing thermal stability by introducing stabilizing mutations. The accurate prediction of the thermodynamic consequences caused by mutations, however, is highly challenging as thermal stability changes are caused by alterations in the free energy of folding. Growing computational power, however, increasingly allows us to use alchemical free energy simulations, such as free energy perturbation or thermodynamic integration, to calculate free energy differences with relatively high accuracy. In this article, we present an automated protocol for setting up alchemical free energy calculations for mutations of naturally occurring amino acids (except for proline) that allows an unprecedented, automated screening of large mutant libraries. To validate the developed protocol, we calculated thermodynamic stability differences for 109 mutations in the microbial Ribonuclease Barnase. The obtained quantitative agreement with experimental data illustrates the potential of the approach in protein engineering and design
New soft-core potential function for molecular dynamics based alchemical free energy calculations.
Full text linkThe fields of rational drug design and protein engineering benefit from accurate free energy calculations based on molecular dynamics simulations. A thermodynamic integration scheme is often used to calculate changes in the free energy of a system by integrating the change of the system’s Hamiltonian with respect to a coupling parameter. These methods exploit nonphysical pathways over thermodynamic cycles involving particle introduction and annihilation. Such alchemical transitions require the modification of the classical nonbonded potential energy terms by applying soft-core potential functions to avoid singularity points. In this work, we propose a novel formulation for a soft-core potential to be applied in nonequilibrium free energy calculations that alleviates singularities, numerical instabilities, and additional minima in the potential energy for all combinations of nonbonded interactions at all intermediate alchemical states. The method was validated by application to (a) the free energy calculations of a closed thermodynamic cycle, (b) the mutation influence on protein thermostability, (c) calculations of small ligand solvation free energies, and (d) the estimation of binding free energies of trypsin inhibitors. The results show that the novel soft-core function provides a robust and accurate general purpose solution to alchemical free energy calculations
Conformational Transitions Upon Ligand Binding: Holo Structure Prediction from Apo Conformations
Full text linkBiological function of proteins is frequently associated with the formation of complexes with small-molecule ligands. Experimental structure determination of such complexes at atomic resolution, however, can be time-consuming and costly. Computational methods for structure prediction of protein/ligand complexes, particularly docking, are as yet restricted by their limited consideration of receptor flexibility, rendering them not applicable for predicting protein/ligand complexes if large conformational changes of the receptor upon ligand binding are involved. Accurate receptor models in the ligand-bound state (holo structures), however, are a prerequisite for successful structure-based drug design. Hence, if only an unbound (apo) structure is available distinct from the ligand-bound conformation, structure-based drug design is severely limited. We present a method to predict the structure of protein/ligand complexes based solely on the apo structure, the ligand and the radius of gyration of the holo structure. The method is applied to ten cases in which proteins undergo structural rearrangements of up to 7.1 A backbone RMSD upon ligand binding. In all cases, receptor models within 1.6 A backbone RMSD to the target were predicted and close-to-native ligand binding poses were obtained for 8 of 10 cases in the top-ranked complex models. A protocol is presented that is expected to enable structure modeling of protein/ligand complexes and structure-based drug design for cases where crystal structures of ligand-bound conformations are not available
Pmx: Automated protein structure and topology generation for alchemical perturbations.
Full text linkComputational protein design requires methods to accurately estimate free energy changes in protein stability or binding upon an amino acid mutation. From the different approaches available, molecular dynamics-based alchemical free energy calculations are unique in their accuracy and solid theoretical basis. The challenge in using these methods lies in the need to generate hybrid structures and topologies representing two physical states of a system. A custom made hybrid topology may prove useful for a particular mutation of interest, however, a high throughput mutation analysis calls for a more general approach. In this work, we present an automated procedure to generate hybrid structures and topologies for the amino acid mutations in all commonly used force fields. The described software is compatible with the Gromacs simulation package. The mutation libraries are readily supported for five force fields, namely Amber99SB, Amber99SB*-ILDN, OPLS-AA/L, Charmm22*, and Charmm36
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