qFit 3: Protein and ligand multiconformer modeling for X-ray crystallographic and single-particle cryo-EM density maps

New X-ray crystallography and cryo-electron microscopy (cryo-EM) approaches yield vast amounts of structural data from dynamic proteins and their complexes. Modeling the full conformational ensemble can provide important biological insights, but identifying and modeling an internally consistent set of alternate conformations remains a formidable challenge. qFit efficiently automates this process by generating a parsimonious multiconformer model. We refactored qFit from a distributed application into software that runs efficiently on a small server, desktop, or laptop. We describe the new qFit 3 software and provide some examples. qFit 3 is open-source under the MIT license, and is available at https://github.com/ExcitedStates/qfit-3.0.

[1]  Gilad Haran,et al.  Direct observation of ultrafast large-scale dynamics of an enzyme under turnover conditions , 2018, Proceedings of the National Academy of Sciences.

[2]  Bengt Samuelsson,et al.  Binding of Pro-Gly-Pro at the active site of leukotriene A4 hydrolase/aminopeptidase and development of an epoxide hydrolase selective inhibitor , 2014, Proceedings of the National Academy of Sciences.

[3]  V. Hornak,et al.  Comparison of multiple Amber force fields and development of improved protein backbone parameters , 2006, Proteins.

[4]  J. Richardson,et al.  The penultimate rotamer library , 2000, Proteins.

[5]  Frank von Delft,et al.  Proper modelling of ligand binding requires an ensemble of bound and unbound states , 2017, Acta crystallographica. Section D, Structural biology.

[6]  David A Sivak,et al.  Crystal cryocooling distorts conformational heterogeneity in a model Michaelis complex of DHFR. , 2014, Structure.

[7]  M. Hamberg,et al.  A dynamic Asp–Arg interaction is essential for catalysis in microsomal prostaglandin E2 synthase , 2016, Proceedings of the National Academy of Sciences.

[8]  Po-Nan Li,et al.  Sequence-guided protein structure determination using graph convolutional and recurrent networks , 2020, 2020 IEEE 20th International Conference on Bioinformatics and Bioengineering (BIBE).

[9]  Ankur Dhanik,et al.  Modeling discrete heterogeneity in X-ray diffraction data by fitting multi-conformers. , 2009, Acta crystallographica. Section D, Biological crystallography.

[10]  Paul D Adams,et al.  Modelling dynamics in protein crystal structures by ensemble refinement , 2012, eLife.

[11]  Sebastian Kelm,et al.  A multi-crystal method for extracting obscured crystallographic states from conventionally uninterpretable electron density , 2017, Nature Communications.

[12]  Michael E. Wall,et al.  Mix-and-inject XFEL crystallography reveals gated conformational dynamics during enzyme catalysis , 2019, Proceedings of the National Academy of Sciences.

[13]  Henry van den Bedem,et al.  qFit-ligand Reveals Widespread Conformational Heterogeneity of Drug-Like Molecules in X-Ray Electron Density Maps. , 2018, Journal of medicinal chemistry.

[14]  Nathan S. Babcock,et al.  Model Selection for Biological Crystallography , 2018, bioRxiv.

[15]  Colin A. Smith,et al.  Backrub-like backbone simulation recapitulates natural protein conformational variability and improves mutant side-chain prediction. , 2008, Journal of molecular biology.

[16]  E A Merritt,et al.  Expanding the model: anisotropic displacement parameters in protein structure refinement. , 1999, Acta crystallographica. Section D, Biological crystallography.

[17]  Niels Volkmann,et al.  Confidence intervals for fitting of atomic models into low-resolution densities , 2009, Acta crystallographica. Section D, Biological crystallography.

[18]  J. Holton,et al.  Protein structural ensembles are revealed by redefining X-ray electron density noise , 2013, Proceedings of the National Academy of Sciences.

[19]  J. Richardson,et al.  Asparagine and glutamine: using hydrogen atom contacts in the choice of side-chain amide orientation. , 1999, Journal of molecular biology.

[20]  Ewen Callaway.  Revolutionary cryo-EM is taking over structural biology , 2020, Nature.

[21]  Randy J Read,et al.  Real-space refinement in PHENIX for cryo-EM and crystallography , 2018, bioRxiv.

[22]  Christopher J. Williams,et al.  Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix , 2019, Acta crystallographica. Section D, Structural biology.

[23]  J. Fraser,et al.  Integrative, dynamic structural biology at atomic resolution—it's about time , 2015, Nature Methods.

[24]  Rama Ranganathan,et al.  Electric-field-stimulated protein mechanics , 2016, Nature.

[25]  D. Kern,et al.  Hidden alternate structures of proline isomerase essential for catalysis , 2010 .

[26]  Wen Hwa Lee,et al.  Large-Scale Structural Analysis of the Classical Human Protein Tyrosine Phosphatome , 2009, Cell.

[27]  Ian W. Davis,et al.  The backrub motion: how protein backbone shrugs when a sidechain dances. , 2006, Structure.

[28]  Henry van den Bedem,et al.  Exposing Hidden Alternative Backbone Conformations in X-ray Crystallography Using qFit , 2015, bioRxiv.

[29]  John Beale,et al.  Resolving polymorphs and radiation-driven effects in microcrystals using fixed-target serial synchrotron crystallography , 2018, Acta crystallographica. Section D, Structural biology.

[30]  Hakan Atakisi,et al.  Effects of protein-crystal hydration and temperature on side-chain conformational heterogeneity in monoclinic lysozyme crystals. , 2018, Acta crystallographica. Section D, Structural biology.

[31]  H. Ng,et al.  Automated electron‐density sampling reveals widespread conformational polymorphism in proteins , 2010, Protein science : a publication of the Protein Society.

[32]  Frank von Delft,et al.  An expanded allosteric network in PTP1B by multitemperature crystallography, fragment screening, and covalent tethering , 2018, eLife.

[33]  Takanori Nakane,et al.  Single-particle cryo-EM at atomic resolution , 2020, Nature.

[34]  Garth J. Williams,et al.  Time-resolved serial crystallography captures high-resolution intermediates of photoactive yellow protein , 2014, Science.

[35]  T. N. Bhat,et al.  The Protein Data Bank , 2000, Nucleic Acids Res..

[36]  Jesse B. Hopkins,et al.  Figures and figure supplements Mapping the conformational landscape of a dynamic enzyme by multitemperature and XFEL crystallography , 2016 .

[37]  Ashwin Chari,et al.  Breaking the next Cryo-EM resolution barrier – Atomic resolution determination of proteins! , 2020, bioRxiv.

[38]  Xianglei Zhang,et al.  Exploration of Fragment Binding Poses Leading to Efficient Discovery of Highly Potent and Orally Effective Inhibitors of FABP4 for Anti-inflammation. , 2020, Journal of medicinal chemistry.

[39]  Paul D. Adams,et al.  Macromolecular refinement of X-ray and cryo-electron microscopy structures with Phenix / OPLS3e for improved structure and ligand quality , 2020, bioRxiv.

[40]  J. Tainer,et al.  The R-factor gap in macromolecular crystallography: an untapped potential for insights on accurate structures , 2014, The FEBS journal.

[41]  Alexander M. Wolff,et al.  Temperature-Jump Solution X-ray Scattering Reveals Distinct Motions in a Dynamic Enzyme , 2018, bioRxiv.

[42]  Randy J. Read,et al.  Iterative-build OMIT maps: map improvement by iterative model building and refinement without model bias , 2008, Acta crystallographica. Section D, Biological crystallography.

[43]  Daniel Herschlag,et al.  A Robust Method for Collecting X-ray Diffraction Data from Protein Crystals across Physiological Temperatures , 2020, bioRxiv.

[44]  Charlotte M. Deane,et al.  Partial-occupancy binders identified by the Pan-Dataset Density Analysis method offer new chemical opportunities and reveal cryptic binding sites , 2017, Structural dynamics.

[45]  Itay Lotan,et al.  Real-space protein-model completion: an inverse-kinematics approach. , 2005, Acta crystallographica. Section D, Biological crystallography.

[46]  Mark A Hallen,et al.  Dead‐end elimination with perturbations (DEEPer): A provable protein design algorithm with continuous sidechain and backbone flexibility , 2013, Proteins.

[47]  David Baker,et al.  PROTEINS: Structure, Function, and Bioinformatics 58:893–904 (2005) A “Solvated Rotamer ” Approach to Modeling Water- Mediated Hydrogen Bonds at Protein–Protein Interfaces , 2022 .

[48]  Paul D. Adams,et al.  Polder maps: improving OMIT maps by excluding bulk solvent , 2017, Acta crystallographica. Section D, Structural biology.

[49]  Nathaniel Echols,et al.  Accessing protein conformational ensembles using room-temperature X-ray crystallography , 2011, Proceedings of the National Academy of Sciences.

[50]  Henry van den Bedem,et al.  Conformational variation of proteins at room temperature is not dominated by radiation damage , 2017, Journal of synchrotron radiation.

[51]  Ezequiel Panepucci,et al.  EIGER detector: application in macromolecular crystallography , 2016, Acta crystallographica. Section D, Structural biology.