Introduction

The molecular dynamics flexible fitting (MDFF) method can be used to flexibly fit atomic structures into density maps. The method was originally described in the manuscript:

Flexible fitting of atomic structures into electron microscopy maps using molecular dynamics. Leonardo G. Trabuco$^1$ , Elizabeth Villa$^1$ , Kakoli Mitra, Joachim Frank, and Klaus Schulten. Structure, 16:673-683, 2008.

The MDFF method can also be used to refine structures using low-resolution X-ray crystallography. This method, xMDFF was originally described in the manuscript:

xMDFF: Molecular dynamics flexible fitting of low-resolution X-Ray structures. Ryan McGreevy$^1$ , Abhishek Singharoy$^1$ , Qufei Li, Jingfen Zhang, Dong Xu, Eduardo Perozo, and Klaus Schulten. Acta Crystallographica D70, 2344-2355, 2014.

We recommend reading the following practical guides before going through this tutorial:

Molecular dynamics flexible fitting: A practical guide to combine cryo-electron microscopy and x-ray crystallography. Leonardo G. Trabuco$^1$ , Elizabeth Villa$^1$ , Eduard Schreiner, Christopher B. Harrison, and Klaus Schulten. Methods, 49:174-180, 2009.

Advances in the molecular dynamics flexible fitting method for cryo-EM modeling. Ryan McGreevy, Ivan Teo, Abhishek Singharoy, and Klaus Schulten. Methods, 100, 50-60, 2016.

Required software

The necessary capabilities for setting up and analyzing MDFF simulations are implemented in VMD (Visual Molecular Dynamics), a molecular visualization and analysis program. MDFF simulations are performed using NAMD (NAnoscale Molecular Dynamics), a molecular dynamics simulation program. Both VMD and NAMD are developed by the Theoretical and Computational Biophysics Group at the University of Illinois at Urbana-Champaign.

To apply the MDFF method you need to download and install both VMD and NAMD. Download and installation instructions can be found following the links above (if you are reading the electronic version of this document). In this tutorial we assume you are familiar with VMD; thus, we recommend that you complete the VMD tutorial beforehand. Completing the NAMD tutorial is not critical for understanding this tutorial, but it is nonetheless recommended.

MDFF can also be used to refine structures from low-resolution X-ray crystallography through a method called xMDFF, covered as part of this tutorial. xMDFF does not fit structures into cryo-EM maps, instead using crystallographic maps which must be generated using the PHENIX software suite. Because of this reliance on PHENIX, you will need to have a recent version installed and executable from the command line, which can be obtained from http://www.phenix-online.org/.

Tutorial Topics and Files

The tutorial starts with a brief overview of MDFF commands available in VMD (Section 1). In Section 2, a simple example of MDFF in vacuo is worked out. This first example uses two atomic structures of adenylate kinase in different conformations, and a simulated map is generated from one of the conformations, which is then used as a target for MDFF. All the basic steps for setting up, running, and analyzing MDFF simulations are covered. In Section 3, a similar MDFF simulation is performed, but this time in explicit solvent. In Section 4, use of domain restraints to maintain rigid domain during MDFF simulations is discussed. In Section 5, use of symmetry restraintes for MDFF of symmetric molecules is covered. In Section 6, xMDFF, an adaptation of MDFF for low-resolution X-ray crystallography is demonstrated. In Section 7, use of the MDFF Graphical User Interface (GUI) instead of the command line is demonstrated, along with interactive MDFF which allows users to manually manipulate their structure during an MDFF simulation.

Table 1: Files provided for each section of this tutorial. All files can be found at the mdff-tutorial-files directory.

Section 2: A simple MDFF example

1ake-colores.pdb

1ake-initial.pdb

4ake-target.pdb

adk-step1-result.dcd

adk-step2-result.dcd

Section 3: MDFF with explicit solvent

adk-solvent-step1-result.dcd

adk-solvent-step2-result.dcd

Section 4: MDFF with Domain Restraints

acoasyn-initial.pdb

acoasyn-initial.psf

acoasyn-target.dx

acoasyn-target.pdb

domain-step1-result.dcd

no-domain-step1-result.dcd

par_all27_prot_lipid_na.inp

Section 5: MDFF with Symmetry Restraints

helix.pdb

helix.psf

helix-target.dx

helix-matrices.txt

no-symmetry-step1-result.dcd

par_all27_prot_lipid_na.inp

set_symmetry.tcl

symmetry-step1-result.dcd