“Conditional Restraints”: Restraining the Free Atoms in ARP/wARP

Wijnand T.M Mooij,Garib N Murshudov,Krista Joosten,Serge X Cohen,Anastassis Perrakis

doi:10.1016/j.str.2008.12.011

Abstract

SummaryThe automated building of a protein model into an electron density map remains a challenging problem. In the ARP/wARP approach, model building is facilitated by initially interpreting a density map with free atoms of unknown chemical identity; all structural information for such chemically unassigned atoms is discarded. Here, this is remedied by applying restraints between free atoms, and between free atoms and a partial protein model. These are based on geometric considerations of protein structure and tentative (conditional) assignments for the free atoms. Restraints are applied in the REFMAC5 refinement program and are generated on an ad hoc basis, allowing them to fluctuate from step to step. A large set of experimentally phased and molecular replacement structures showcases individual structures where automated building is improved drastically by the conditional restraints. The concept and implementation we present can also find application in restraining geometries, such as hydrogen bonds, in low-resolution refinement.

Highlights

The determination of a protein structure by X-ray crystallography is a process that involves many different stages, from protein production, crystallization, and data collection, to structure solution and refinement
Computational Methods The ARP/wARP workflow has been described previously (Morris et al, 2003). It consists of a number of tasks that are performed in an iterative fashion, and briefly outlined as follows: Start: The procedure typically starts by building a model of free atoms in an electron density map
Atoms with a low electron density value in the 2mFo-DFc map or too far from all other atoms are removed; free atoms are added at positions of high density in the mFo-DFc map, provided they are close to existing atoms

Summary

Introduction

The determination of a protein structure by X-ray crystallography is a process that involves many different stages, from protein production, crystallization, and data collection, to structure solution and refinement. Free atoms serve two reasons in the ARP/wARP formulation: first, they are used to obtain better electron density maps through refinement; second, during model building they serve both as possible Ca guides for the protein backbone and as guides for the sequence assignment. It becomes difficult to refine the free atoms or hybrid model structure with the limited number of experimental observations and without chemical restraints.

Results

Conclusion