Residue contacts predicted by evolutionary covariance extend the application of ab initio molecular replacement to larger and more challenging protein folds.

Felix Simkovic,Olga Mayans,Martyn D Winn,Jens M H Thomas,Daniel J Rigden,Ronan M Keegan

doi:10.1107/s2052252516008113

Abstract

For many protein families, the deluge of new sequence information together with new statistical protocols now allow the accurate prediction of contacting residues from sequence information alone. This offers the possibility of more accurate ab initio (non-homology-based) structure prediction. Such models can be used in structure solution by molecular replacement (MR) where the target fold is novel or is only distantly related to known structures. Here, AMPLE, an MR pipeline that assembles search-model ensembles from ab initio structure predictions ('decoys'), is employed to assess the value of contact-assisted ab initio models to the crystallographer. It is demonstrated that evolutionary covariance-derived residue-residue contact predictions improve the quality of ab initio models and, consequently, the success rate of MR using search models derived from them. For targets containing β-structure, decoy quality and MR performance were further improved by the use of a β-strand contact-filtering protocol. Such contact-guided decoys achieved 14 structure solutions from 21 attempted protein targets, compared with nine for simple Rosetta decoys. Previously encountered limitations were superseded in two key respects. Firstly, much larger targets of up to 221 residues in length were solved, which is far larger than the previously benchmarked threshold of 120 residues. Secondly, contact-guided decoys significantly improved success with β-sheet-rich proteins. Overall, the improved performance of contact-guided decoys suggests that MR is now applicable to a significantly wider range of protein targets than were previously tractable, and points to a direct benefit to structural biology from the recent remarkable advances in sequencing.

Highlights

Molecular replacement (MR) is the most common technique for deriving the lost phase information of the unknown target structure in X-ray crystallography
Such sources include ideal secondary-structure elements or structural motifs (Rodrıguez et al, 2009), libraries of tertiary structural cores derived from mining the Protein Data Bank (PDB; Sammito et al, 2013) and ab initio protein structure predictions (‘decoys’; Qian et al, 2007; Rigden et al, 2008; Das & Baker, 2009; Bibby et al, 2012; Keegan et al, 2015; Ramisch et al, 2015; Thomas et al, 2015)
Fragments were picked for unassisted Rosetta modelling with secondary-structure prediction from PSIPRED (McGuffin et al, 2000) and for contact-assisted decoys with the secondary-structure prediction obtained during evolutionary covariance analysis

Summary

Introduction

Molecular replacement (MR) is the most common technique for deriving the lost phase information of the unknown target structure in X-ray crystallography. Since the successful separation of direct and indirect contacts, various evolutionary covariance-analysis applications have been developed to increase the accuracy and speed of contact predictions (Balakrishnan et al, 2011; Jeong & Kim, 2012; Jones et al, 2012; Ekeberg et al, 2013, 2014; Kamisetty et al, 2013; Wang & Xu, 2013; Feinauer et al, 2014; Kajan et al, 2014; Schneider & Brock, 2014; Seemayer et al, 2014; Skwark et al, 2014) These applications can be divided into three categories depending on the cooperative statistical model implemented to derive evolutionary covariance amongst multiple homologous sequences. We report that combining independently obtained contact maps further improves decoy quality, which in turn extends the tractable MR target range to -rich proteins

Data set

Evolutionary covariance analysis

Conversion of contact maps to contact restraints

Ab initio structure prediction of decoys

Molecular replacement

Results