Computer Physics Communications 185 (2014) 2920–2929 Contents lists available at ScienceDirect Computer Physics Communications journal homepage: www.elsevier.com/locate/cpc GMXPBSA 2.0: A GROMACS tool to perform MM/PBSA and computational alanine scanningI C. Paissoni a, D. Spiliotopoulos a,1, G. Musco a, A. Spitaleri a,b,∗ a Biomolecular NMR Unit, S. Raffaele Scientific Institute, via Olgettina 58, Milan 20132, Italy b Drug Discovery and Development, Istituto Italiano di Tecnologia, Via Morego 30, Genoa 16163, Italy article info a b s t r a c t Article history: GMXPBSA 2.0 is a user-friendly suite of Bash/Perl scripts for streamlining MM/PBSA calculations on Received 9 January 2014 structural ensembles derived from GROMACS trajectories, to automatically calculate binding free energies Received in revised form for protein–protein or ligand–protein complexes. GMXPBSA 2.0 is flexible and can easily be customized 19 May 2014 to specific needs. Additionally, it performs computational alanine scanning (CAS) to study the effects Accepted 17 June 2014 of ligand and/or receptor alanine mutations on the free energy of binding. Calculations require only Available online 2 July 2014 for protein–protein or protein–ligand MD simulations. GMXPBSA 2.0 performs different comparative analysis, including a posteriori generation of alanine mutants of the wild-type complex, calculation of Keywords: Molecular dynamics simulation the binding free energy values of the mutant complexes and comparison of the results with the wild-type Binding free energy system. Moreover, it compares the binding free energy of different complexes trajectories, allowing the Virtual screening study the effects of non-alanine mutations, post-translational modifications or unnatural amino acids on GROMACS the binding free energy of the system under investigation. Finally, it can calculate and rank relative affinity Computational alanine scanning to the same receptor utilizing MD simulations of proteins in complex with different ligands. In order to MM/PBSA dissect the different MM/PBSA energy contributions, including molecular mechanic (MM), electrostatic contribution to solvation (PB) and nonpolar contribution to solvation (SA), the tool combines two freely available programs: the MD simulations software GROMACS and the Poisson–Boltzmann equation solver APBS. All the calculations can be performed in single or distributed automatic fashion on a cluster facility in order to increase the calculation by dividing frames across the available processors. The program is freely available under the GPL license. Program summary Program title: GMXPBSA 2.0 Catalogue identifier: AETQ_v1_0 Program summary URL: http://cpc.cs.qub.ac.uk/summaries/AETQ_v1_0.html Program obtainable from: CPC Program Library, Queen's University, Belfast, N. Ireland Licensing provisions: GNU General Public License, version 3 No. of lines in distributed program, including test data, etc.: 185 937 No. of bytes in distributed program, including test data, etc.: 7 074 217 Distribution format: tar.gz Programming language: Bash, Perl. Computer: Any computer. Operating system: Linux, Unix OS. RAM: ∼2 GB I This paper and its associated computer program are available via the Computer Physics Communication homepage on ScienceDirect (http://www.sciencedirect.com/ science/journal/00104655). ∗ Corresponding author at: Drug Discovery and Development, Istituto Italiano di Tecnologia, Via Morego 30, Genoa 16163, Italy. Tel.: +39 3485188790. E-mail address: [email protected] (A. Spitaleri). 1 Present address: Computational Structural Biology Biochemisches Institut Universität Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland. http://dx.doi.org/10.1016/j.cpc.2014.06.019 0010-4655/' 2014 Elsevier B.V. All rights reserved. C. Paissoni et al. / Computer Physics Communications 185 (2014) 2920–2929 2921 Classification: 3. External routines: APBS (http://www.Poissonboltzmann.org/apbs/) and GROMACS installations (http:// www.gromacs.org). Optionally LaTeX. Nature of problem: Calculates the Molecular Mechanics (MM) data (Lennard-Jones and Coulomb terms) and the solvation energy terms (polar and nonpolar terms respectively) from an ensemble of structures derived from GROMACS molecular dynamics simulation trajectory. These calculations are performed for each single component of the simulated complex, including protein and ligand. In order to cancel out artefacts an identical grid setup for each component, including complex, protein and ligand, is required. Performs statistical analysis on the extracted data and comparison with wild-type complex in case of either computational alanine scanning or calculations on a set of simulations. Evaluates possible outliers in the frames extracted from the simulations during the binding free energy calculations. Solution method: The tool combines the freely available programs GROMACS and APBS to: 1. extract frames from a single or multiple complex molecular dynamics (MD) simulation, allowing comparison between multiple trajectories; 2. split the complex frames in the single components including complex, protein and ligand; 3. calculate the Lennard-Jones and Coulomb energy values (MM terms); 4. calculate the polar solvation energy values using the implicit solvation Poisson–Boltzmann model (PB); 5. calculate the nonpolar solvation energy value based on the solvent accessible surface area (SASA); 6. combine all the calculated terms into the final binding free energy value; 7. repeat the same procedure from point 1 to 6 for each simulation in case of computational alanine scanning (CAS) or simultaneous comparison of different MDs. Restrictions: Input format files compatible with GROMACS engine 4.5 and later versions. Availability of the force field or of the topology files. Running time: On a single core, Lennard-Jones, Coulomb and nonpolar solvation terms calculations require a few min- utes. The time required for polar solvation terms calculations depends on the system size. ' 2014 Elsevier B.V. All rights reserved. 1. Introduction where Eint indicates bond, angle, and torsional angle energies, and Ecoul and ELJ denote the intramolecular electrostatic and Lennard- MM/PBSA is a versatile method to calculate the binding free en- Jones energies, respectively. ergies of a protein–ligand complex [1]. It incorporates the effects The solvation term Gsolv in Eq. (4) is split into polar Gpolar and of thermal averaging with a force field/continuum solvent model nonpolar contributions, Gnonpolar: to post-process a series of representative snapshots from MD trajectories. MM/PBSA has been successfully applied to compute Gsolv D Gpolar C Gnonpolar (4) the binding free energy of numerous protein–ligand interactions GMXPBSA 2.0 calculates G and G with Adaptive Pois- [2–5]. The method expresses the free energy of binding as the dif- polar nonpolar son–Boltzmann Solver (APBS) program [7]. ference between the free energy of the complex and the free energy The polar contribution G refers to the energy required to of the receptor plus the ligand (end-state method). This difference polar transfer the solute from a continuum medium with a low dielectric is averaged over a number of trajectory snapshots [6]. Of note, the constant (" D 1) to a continuum medium with the dielectric con- MM/PBSA approach allows for a rapid estimation of the variation stant of water (" D 80). G is calculated using the non-linearized in the free energy of binding, with the caveat that generally it does polar or linearized Poisson–Boltzmann equation. The nonpolar contribu- not reproduce the absolute binding free energy values. Neverthe- tion G is considered proportional to the solvent accessible less, it usually exhibits good correlations with experiments, thus nonpolar surface area (SASA): representing a fair compromise between efficiency and efficacy for the calculation and comparison of binding free energy variations. Gnonpolar D γ SASA C β (5) The theory underlying MM/PBSA approach has been described pre- − −2 − viously [6]. Briefly, the binding free energy of a protein molecule to where γ D 0:0227 kJ mol 1Å and β D 0 kJ mol 1 [8]. The di- a ligand molecule in solution is defined as: electric boundary is defined using a probe of radius 1.4 Å. Binding free energy calculations based on the MM/PBSA ap- ∆Gbinding D Gcomplex − .Gprotein C Gligand/: (1) proach can be performed either according to the three trajecto- A MD simulation is performed to generate a thermodynamically ries method (TTM) or to the single trajectory method (STM). The weighted ensemble of structures. The free energy term is calcu- TTM requires three separate MD simulations on the three system lated as an average over the considered structures: components including the complex, the free ligand and the free re- ceptor. This is a computationally demanding approach and prone hGi D hEMMi C hGsolvi − T hSMMi: (2) to structural noise [3,5]. Conversely, the STM requires a single tra- The energetic term E is defined as: MM jectory run for the complex, whereby both the protein and ligand EMM D Eint C Ecoul C ELJ (3) structures are extracted directly from the complex structure [3], 2922 C. Paissoni et al. / Computer Physics Communications 185 (2014) 2920–2929 Fig. 1. Workflow diagram for GMXPBSA 2.0. Diagram describing the general GMXPBSA 2.0 workflow scheme. GMXPBSA 2.0 combines the GROMACS and APBS programs in order to use the frames extracted from the molecular dynamics simulations and to calculate the binding free energy. thus zeroing out the Eint term. In this