scholarly journals Exploring the speed and performance of molecular replacement withAMPLEusingQUARK ab initioprotein models

2015 ◽  
Vol 71 (2) ◽  
pp. 338-343 ◽  
Author(s):  
Ronan M. Keegan ◽  
Jaclyn Bibby ◽  
Jens Thomas ◽  
Dong Xu ◽  
Yang Zhang ◽  
...  
Keyword(s):  
Protein Structure ◽  
Ab Initio ◽  
Molecular Replacement ◽  
Test Set ◽  
Search Models ◽  
Structure Solution ◽  
And Performance ◽  
Almost All

AMPLEclusters and truncatesab initioprotein structure predictions, producing search models for molecular replacement. Here, an interesting degree of complementarity is shown between targets solved using the differentab initiomodelling programsQUARKandROSETTA. Search models derived from either program collectively solve almost all of the all-helical targets in the test set. Initial solutions produced byPhaserafter only 5 min perform surprisingly well, improving the prospects forin situstructure solution byAMPLEduring synchrotron visits. Taken together, the results show the potential forAMPLEto run more quickly and successfully solve more targets than previously suspected.

scholarly journals Rapid molecular replacement of coiled-coil and transmembrane proteins with AMPLE

2014 ◽  
Vol 70 (a1) ◽  
pp. C347-C347
Author(s):  
Jens Thomas ◽  
Ronan Keegan ◽  
Jaclyn Bibby ◽  
Martyn Winn ◽  
Olga Mayans ◽  
...  
Keyword(s):  
Ab Initio ◽  
Protein Structures ◽  
Coiled Coil ◽  
Molecular Replacement ◽  
Helical Structures ◽  
Desktop Computer ◽  
Search Models ◽  
Nmr Structures ◽  
Structure Solution ◽  
Helical Geometry

Molecular Replacement (MR) is an increasingly popular route to protein structure solution. AMPLE[1] is a software pipeline that uses either cheaply obtained ab inito protein models, or NMR structures to extend the scope of MR, allowing it to solve entirely novel protein structures in a completely automated pipeline on a standard desktop computer. AMPLE employs a cluster-and-truncate approach, combined with multiple modes of side chain treatment, to analyse the candidate models and extract the consensual features most likely to solve the structure. The search models generated in this way are screened by MrBump using Phaser and Molrep and correct solutions are detected using main chain tracing and phase modification with Shelxe. AMPLE proved capable of processing rapidly obtained ab initio structure predictions into successful search models and more recently proved effective in assembling NMR structures for MR[2]. Coiled-coil proteins are a distinct class of protein fold whose structure solution by MR is not typically straightforward. We show here that AMPLE can quickly and routinely solve most coiled-coil structures using ab initio predictions from Rosetta. The predictions are generally not globally accurate, but by encompassing different degrees of truncation of clustered models, AMPLE succeeds by sampling across a range of search models. These sometimes succeed through capturing locally well-modelled conformations, but often simply contain small helical units. Remarkably, the latter regularly succeed despite out-of-register placement and poor MR statistics. We demonstrate that single structures derived from successful ensembles perform less well, and comparable ideal helices solve few targets. Thus, both modelling of distortions from ideal helical geometry and the ensemble nature of the search models contribute to success. AMPLE is a framework applicable to any set of input structures in which variability is correlated with inaccuracy. We also present preliminary data demonstrating structure solution of transmembrane helical structures using Rosetta modelling. We finally consider future sources of starting models which offer the hope that MR with AMPLE, in the absence of close homology between a known structure and the target, may soon be possible with larger proteins.

scholarly journals Application of theAMPLEcluster-and-truncate approach to NMR structures for molecular replacement

2013 ◽  
Vol 69 (11) ◽  
pp. 2194-2201 ◽  
Author(s):  
Jaclyn Bibby ◽  
Ronan M. Keegan ◽  
Olga Mayans ◽  
Martyn D. Winn ◽  
Daniel J. Rigden
Keyword(s):  
Protein Structure ◽  
Ab Initio ◽  
Target Protein ◽  
Molecular Replacement ◽  
Search Models ◽  
Sequence Identity ◽  
Nmr Structures ◽  
Core Cluster ◽  
Structural Divergence ◽  
Improved Performance

AMPLEis a program developed for clustering and truncatingab initioprotein structure predictions into search models for molecular replacement. Here, it is shown that its core cluster-and-truncate methods also work well for processing NMR ensembles into search models.Rosettaremodelling helps to extend success to NMR structures bearing low sequence identity or high structural divergence from the target protein. Potential future routes to improved performance are considered and practical, general guidelines on usingAMPLEare provided.

scholarly journals Approaches toab initiomolecular replacement of α-helical transmembrane proteins

2017 ◽  
Vol 73 (12) ◽  
pp. 985-996 ◽  
Author(s):  
Jens M. H. Thomas ◽  
Felix Simkovic ◽  
Ronan Keegan ◽  
Olga Mayans ◽  
Chengxin Zhang ◽  
...  
Keyword(s):  
Ab Initio ◽  
Transmembrane Protein ◽  
Protein Structures ◽  
Transmembrane Proteins ◽  
Crystallographic Structure ◽  
Molecular Replacement ◽  
Search Models ◽  
Contact Information ◽  
Structure Solution ◽  
The Ideal

α-Helical transmembrane proteins are a ubiquitous and important class of proteins, but present difficulties for crystallographic structure solution. Here, the effectiveness of theAMPLEmolecular replacement pipeline in solving α-helical transmembrane-protein structures is assessed using a small library of eight ideal helices, as well as search models derived fromab initiomodels generated both with and without evolutionary contact information. The ideal helices prove to be surprisingly effective at solving higher resolution structures, butab initio-derived search models are able to solve structures that could not be solved with the ideal helices. The addition of evolutionary contact information results in a marked improvement in the modelling and makes additional solutions possible.

Error-estimation-guided rebuilding ofde novomodels increases the success rate ofab initiophasing

2012 ◽  
Vol 68 (11) ◽  
pp. 1522-1534 ◽  
Author(s):  
Rojan Shrestha ◽  
David Simoncini ◽  
Kam Y. J. Zhang
Keyword(s):  
Protein Structure ◽  
Ab Initio ◽  
Diffraction Data ◽  
Structure Prediction ◽  
De Novo ◽  
Coarse Grained ◽  
Data Sets ◽  
Molecular Replacement ◽  
High Quality ◽  
Protein Targets

Recent advancements in computational methods for protein-structure prediction have made it possible to generate the high-qualityde novomodels required forab initiophasing of crystallographic diffraction data using molecular replacement. Despite those encouraging achievements inab initiophasing usingde novomodels, its success is limited only to those targets for which high-qualityde novomodels can be generated. In order to increase the scope of targets to whichab initiophasing withde novomodels can be successfully applied, it is necessary to reduce the errors in thede novomodels that are used as templates for molecular replacement. Here, an approach is introduced that can identify and rebuild the residues with larger errors, which subsequently reduces the overall Cαroot-mean-square deviation (CA-RMSD) from the native protein structure. The error in a predicted model is estimated from the average pairwise geometric distance per residue computed among selected lowest energy coarse-grained models. This score is subsequently employed to guide a rebuilding process that focuses on more error-prone residues in the coarse-grained models. This rebuilding methodology has been tested on ten protein targets that were unsuccessful using previous methods. The average CA-RMSD of the coarse-grained models was improved from 4.93 to 4.06 Å. For those models with CA-RMSD less than 3.0 Å, the average CA-RMSD was improved from 3.38 to 2.60 Å. These rebuilt coarse-grained models were then converted into all-atom models and refined to produce improvedde novomodels for molecular replacement. Seven diffraction data sets were successfully phased using rebuiltde novomodels, indicating the improved quality of these rebuiltde novomodels and the effectiveness of the rebuilding process. Software implementing this method, calledMORPHEUS, can be downloaded from http://www.riken.jp/zhangiru/software.html.

scholarly journals Crystallographic ab initio protein structure solution below atomic resolution

2009 ◽  
Vol 6 (9) ◽  
pp. 651-653 ◽  
Author(s):  
Dayté D Rodríguez ◽  
Christian Grosse ◽  
Sebastian Himmel ◽  
César González ◽  
Iñaki M de Ilarduya ◽  
...  
Keyword(s):  
Protein Structure ◽  
Ab Initio ◽  
Atomic Resolution ◽  
Structure Solution

scholarly journals Crystallographic molecular replacement using an in silico-generated search model of SARS-CoV-2 ORF8

2021 ◽  
Author(s):  
Thomas G. Flower ◽  
James H. Hurley
Keyword(s):  
Ab Initio ◽  
Structural Parameters ◽  
Search Model ◽  
Problem Solution ◽  
Molecular Replacement ◽  
Phase Problem ◽  
Search Models ◽  
Sad Phasing ◽  
Predicted Model ◽  
Β Sheet

AbstractThe majority of crystal structures are determined by the method of molecular replacement (MR). The range of application of MR is limited mainly by the need for an accurate search model. In most cases, pre-existing experimentally determined structures are used as search models. In favorable cases, ab initio predicted structures have yielded search models adequate for molecular replacement. The ORF8 protein of SARS-CoV-2 represents a challenging case for MR using an ab initio prediction because ORF8 has an all β-sheet fold and few orthologs. We previously determined experimentally the structure of ORF8 using the single anomalous dispersion (SAD) phasing method, having been unable to find an MR solution to the crystallographic phase problem. Following a report of an accurate prediction of the ORF8 structure, we assessed whether the predicted model would have succeeded as an MR search model. A phase problem solution was found, and the resulting structure was refined, yielding structural parameters equivalent to the original experimental solution.

scholarly journals Routine phasing of coiled-coil protein crystal structures withAMPLE

IUCrJ ◽  
2015 ◽  
Vol 2 (2) ◽  
pp. 198-206 ◽  
Author(s):  
Jens M. H. Thomas ◽  
Ronan M. Keegan ◽  
Jaclyn Bibby ◽  
Martyn D. Winn ◽  
Olga Mayans ◽  
...  
Keyword(s):  
Ab Initio ◽  
Crystal Structures ◽  
Coiled Coil ◽  
Protein Crystal ◽  
Molecular Replacement ◽  
Search Models ◽  
Homologous Protein ◽  
Coil Structure ◽  
Convenient Route ◽  
Chain Lengths

Coiled-coil protein folds are among the most abundant in nature. These folds consist of long wound α-helices and are architecturally simple, but paradoxically their crystallographic structures are notoriously difficult to solve with molecular-replacement techniques. The programAMPLEcan solve crystal structures by molecular replacement usingab initiosearch models in the absence of an existent homologous protein structure.AMPLEhas been benchmarked on a large and diverse test set of coiled-coil crystal structures and has been found to solve 80% of all cases. Successes included structures with chain lengths of up to 253 residues and resolutions down to 2.9 Å, considerably extending the limits on size and resolution that are typically tractable byab initiomethodologies. The structures of two macromolecular complexes, one including DNA, were also successfully solved using their coiled-coil components. It is demonstrated that both theab initiomodelling and the use of ensemble search models contribute to the success ofAMPLEby comparison with phasing attempts using single structures or ideal polyalanine helices. These successes suggest that molecular replacement withAMPLEshould be the method of choice for the crystallographic elucidation of a coiled-coil structure. Furthermore,AMPLEmay be able to exploit the presence of a coiled coil in a complex to provide a convenient route for phasing.

scholarly journals Error estimation guided rebuilding of de novo models for ab initio phasing

2014 ◽  
Vol 70 (a1) ◽  
pp. C1448-C1448
Author(s):  
Rojan Shrestha ◽  
David Simoncini ◽  
Kam Zhang
Keyword(s):  
Protein Structure ◽  
Ab Initio ◽  
Structure Prediction ◽  
De Novo ◽  
Coarse Grained ◽  
Molecular Replacement ◽  
High Quality ◽  
Protein Targets ◽  
Current State

Recent advancement in computational methods for protein structure prediction has made it possible to generate high quality de novo models required for ab initio phasing of crystallographic diffraction data using molecular replacement. Despite those encouraging achievements in ab initio phasing using de novo models, its success is limited only to those targets for which high quality de novo models can be generated. Here, an approach is introduced that can identify and rebuild the residues with larger errors, which subsequently reduces the overall C-alpha root mean square deviations (CA-RMSD) to the native protein structure. The error in a predicted model is estimated by the average pairwise geometric distance per residue computed among selected lowest energy coarse-grained models. This score is subsequently employed to guide a rebuilding process that focuses on more error-prone residues in the coarse-grained models. These rebuilt coarse-grained models were then turned into all-atom models and refined to produce improved de novo models for molecular replacement. This rebuilding methodology has been tested on ten protein targets that were unsuccessful with the current state-of-the-art methods. Seven diffraction datasets were successfully phased using rebuilt de novo models indicating the improved quality of these rebuilt de novo models and the effectiveness of this rebuilding process.

Crystal structure determination and refinementviaSIR2014

2015 ◽  
Vol 48 (1) ◽  
pp. 306-309 ◽  
Author(s):  
Maria Cristina Burla ◽  
Rocco Caliandro ◽  
Benedetta Carrozzini ◽  
Giovanni Luca Cascarano ◽  
Corrado Cuocci ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Ab Initio ◽  
Structure Refinement ◽  
Three Dimensional ◽  
Direct Methods ◽  
Molecular Replacement ◽  
Dimensional Electron ◽  
Large Structures ◽  
Density Maps ◽  
Structure Solution

SIR2014is the latest program of theSIRsuite for crystal structure solution of small, medium and large structures. A variety of phasing algorithms have been implemented, bothab initio(standard or modern direct methods, Patterson techniques,Vive la Différence) and non-ab initio(simulated annealing, molecular replacement). The program contains tools for crystal structure refinement and for the study of three-dimensional electron-density mapsviasuitable viewers.

scholarly journals Extending the scope of coiled-coil crystal structure solution by AMPLE through improved ab initio modelling

2020 ◽  
Vol 76 (3) ◽  
pp. 272-284 ◽  
Author(s):  
Jens M. H. Thomas ◽  
Ronan M. Keegan ◽  
Daniel J. Rigden ◽  
Owen R. Davies
Keyword(s):  
Ab Initio ◽  
Coiled Coil ◽  
Coiled Coils ◽  
Molecular Replacement ◽  
Helical Structures ◽  
Major Barrier ◽  
X Ray Crystallography ◽  
Structure Solution ◽  
Coil Structure ◽  
Experimental Phasing

The phase problem remains a major barrier to overcome in protein structure solution by X-ray crystallography. In recent years, new molecular-replacement approaches using ab initio models and ideal secondary-structure components have greatly contributed to the solution of novel structures in the absence of clear homologues in the PDB or experimental phasing information. This has been particularly successful for highly α-helical structures, and especially coiled-coils, in which the relatively rigid α-helices provide very useful molecular-replacement fragments. This has been seen within the program AMPLE, which uses clustered and truncated ensembles of numerous ab initio models in structure solution, and is already accomplished for α-helical and coiled-coil structures. Here, an expansion in the scope of coiled-coil structure solution by AMPLE is reported, which has been achieved through general improvements in the pipeline, the removal of tNCS correction in molecular replacement and two improved methods for ab initio modelling. Of the latter improvements, enforcing the modelling of elongated helices overcame the bias towards globular folds and provided a rapid method (equivalent to the time requirements of the existing modelling procedures in AMPLE) for enhanced solution. Further, the modelling of two-, three- and four-helical oligomeric coiled-coils, and the use of full/partial oligomers in molecular replacement, provided additional success in difficult and lower resolution cases. Together, these approaches have enabled the solution of a number of parallel/antiparallel dimeric, trimeric and tetrameric coiled-coils at resolutions as low as 3.3 Å, and have thus overcome previous limitations in AMPLE and provided a new functionality in coiled-coil structure solution at lower resolutions. These new approaches have been incorporated into a new release of AMPLE in which automated elongated monomer and oligomer modelling may be activated by selecting `coiled-coil' mode.

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