skip to main content
DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Real-space refinement in PHENIX for cryo-EM and crystallography

Abstract

This article describes the implementation of real-space refinement in thephenix.real_space_refineprogram from thePHENIXsuite. The use of a simplified refinement target function enables very fast calculation, which in turn makes it possible to identify optimal data-restraint weights as part of routine refinements with little runtime cost. Refinement of atomic models against low-resolution data benefits from the inclusion of as much additional information as is available. In addition to standard restraints on covalent geometry,phenix.real_space_refinemakes use of extra information such as secondary-structure and rotamer-specific restraints, as well as restraints or constraints on internal molecular symmetry. The re-refinement of 385 cryo-EM-derived models available in the Protein Data Bank at resolutions of 6 Å or better shows significant improvement of the models and of the fit of these models to the target maps.

Authors:
ORCiD logo [1]; ORCiD logo [2]; ORCiD logo [3];  [2]; ORCiD logo [4];  [5];  [6]
  1. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Shanghai Univ. (China). Dept. of Physics and International Centre for Quantum and Molecular Structures
  2. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division
  3. Univ. of Cambridge (United Kingdom). Cambridge Inst. for Medical Research
  4. Los Alamos National Lab. (LANL), Los Alamos, NM (United States); New Mexico Consortium (NMC), Los Alamos, NM (United States)
  5. Univ. of Lorraine, Vandoeuvre-les-Nancy (France). Faculty of Science and Technology; Inst. of Genetics and Molecular and Cell Biology (IGBMC), CNRS-INSERM-UdS, Illkirch (France). Centre for Integrative Biology
  6. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Univ. of California, Berkeley, CA (United States). Dept. of Bioengineering
Publication Date:
Research Org.:
Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
National Institutes of Health (NIH); French Infrastructure for Integrated Structural Biology (FRISBI); Wellcome Trust, London (United Kingdom); USDOE Office of Science (SC)
OSTI Identifier:
1439491
Alternate Identifier(s):
OSTI ID: 1441336; OSTI ID: 1461166
Report Number(s):
LA-UR-18-20271
Journal ID: ISSN 2059-7983; ACSDAD; PII: S2059798318006551
Grant/Contract Number:  
AC52-06NA25396; GM063210; AC02-05CH11231; 082961/ Z/07/Z; ANR-10-INBS-05
Resource Type:
Published Article
Journal Name:
Acta Crystallographica. Section D. Structural Biology
Additional Journal Information:
Journal Name: Acta Crystallographica. Section D. Structural Biology Journal Volume: 74 Journal Issue: 6; Journal ID: ISSN 2059-7983
Publisher:
IUCr
Country of Publication:
United Kingdom
Language:
English
Subject:
97 MATHEMATICS AND COMPUTING; 59 BASIC BIOLOGICAL SCIENCES; Biological Science

Citation Formats

Afonine, Pavel V., Poon, Billy K., Read, Randy J., Sobolev, Oleg V., Terwilliger, Thomas C., Urzhumtsev, Alexandre, and Adams, Paul D. Real-space refinement in PHENIX for cryo-EM and crystallography. United Kingdom: N. p., 2018. Web. doi:10.1107/S2059798318006551.
Afonine, Pavel V., Poon, Billy K., Read, Randy J., Sobolev, Oleg V., Terwilliger, Thomas C., Urzhumtsev, Alexandre, & Adams, Paul D. Real-space refinement in PHENIX for cryo-EM and crystallography. United Kingdom. doi:10.1107/S2059798318006551.
Afonine, Pavel V., Poon, Billy K., Read, Randy J., Sobolev, Oleg V., Terwilliger, Thomas C., Urzhumtsev, Alexandre, and Adams, Paul D. Fri . "Real-space refinement in PHENIX for cryo-EM and crystallography". United Kingdom. doi:10.1107/S2059798318006551.
@article{osti_1439491,
title = {Real-space refinement in PHENIX for cryo-EM and crystallography},
author = {Afonine, Pavel V. and Poon, Billy K. and Read, Randy J. and Sobolev, Oleg V. and Terwilliger, Thomas C. and Urzhumtsev, Alexandre and Adams, Paul D.},
abstractNote = {This article describes the implementation of real-space refinement in thephenix.real_space_refineprogram from thePHENIXsuite. The use of a simplified refinement target function enables very fast calculation, which in turn makes it possible to identify optimal data-restraint weights as part of routine refinements with little runtime cost. Refinement of atomic models against low-resolution data benefits from the inclusion of as much additional information as is available. In addition to standard restraints on covalent geometry,phenix.real_space_refinemakes use of extra information such as secondary-structure and rotamer-specific restraints, as well as restraints or constraints on internal molecular symmetry. The re-refinement of 385 cryo-EM-derived models available in the Protein Data Bank at resolutions of 6 Å or better shows significant improvement of the models and of the fit of these models to the target maps.},
doi = {10.1107/S2059798318006551},
journal = {Acta Crystallographica. Section D. Structural Biology},
number = 6,
volume = 74,
place = {United Kingdom},
year = {2018},
month = {6}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
DOI: 10.1107/S2059798318006551

Citation Metrics:
Cited by: 53 works
Citation information provided by
Web of Science

Save / Share:

Works referenced in this record:

Structure refinement: some background theory and practical strategies
journal, April 2008


On the limited memory BFGS method for large scale optimization
journal, August 1989

  • Liu, Dong C.; Nocedal, Jorge
  • Mathematical Programming, Vol. 45, Issue 1-3
  • DOI: 10.1007/BF01589116

Improved metrics for comparing structures of macromolecular assemblies determined by 3D electron-microscopy
journal, July 2017

  • Joseph, Agnel Praveen; Lagerstedt, Ingvar; Patwardhan, Ardan
  • Journal of Structural Biology, Vol. 199, Issue 1
  • DOI: 10.1016/j.jsb.2017.05.007

Robust Parameterization of Elastic and Absorptive Electron Atomic Scattering Factors
journal, March 1996

  • Peng, L. M.; Ren, G.; Dudarev, S. L.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 52, Issue 2
  • DOI: 10.1107/S0108767395014371

CryoEM Structure of an Influenza Virus Receptor-Binding Site Antibody–Antigen Interface
journal, June 2017


TRPV1 structures in nanodiscs reveal mechanisms of ligand and lipid action
journal, May 2016


Structural basis for anion conduction in the calcium-activated chloride channel TMEM16A
journal, May 2017


Mechanism of ribosome rescue by ArfA and RF2
journal, March 2017


Atomic-accuracy models from 4.5-Å cryo-electron microscopy data with density-guided iterative local refinement
journal, February 2015

  • DiMaio, Frank; Song, Yifan; Li, Xueming
  • Nature Methods, Vol. 12, Issue 4
  • DOI: 10.1038/nmeth.3286

Flexible torsion-angle noncrystallographic symmetry restraints for improved macromolecular structure refinement
journal, April 2014

  • Headd, Jeffrey J.; Echols, Nathaniel; Afonine, Pavel V.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 70, Issue 5
  • DOI: 10.1107/S1399004714003277

A New Generation of Crystallographic Validation Tools for the Protein Data Bank
journal, October 2011


EMDep: a web-based system for the deposition and validation of high-resolution electron microscopy macromolecular structural information
journal, October 2003


Accurate model annotation of a near-atomic resolution cryo-EM map
journal, March 2017

  • Hryc, Corey F.; Chen, Dong-Hua; Afonine, Pavel V.
  • Proceedings of the National Academy of Sciences, Vol. 114, Issue 12
  • DOI: 10.1073/pnas.1621152114

MolProbity: More and better reference data for improved all-atom structure validation: PROTEIN SCIENCE.ORG
journal, November 2017

  • Williams, Christopher J.; Headd, Jeffrey J.; Moriarty, Nigel W.
  • Protein Science, Vol. 27, Issue 1
  • DOI: 10.1002/pro.3330

Crystallography & NMR System: A New Software Suite for Macromolecular Structure Determination
journal, September 1998

  • Brünger, A. T.; Adams, P. D.; Clore, G. M.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 54, Issue 5
  • DOI: 10.1107/S0907444998003254

Towards automated crystallographic structure refinement with phenix.refine
journal, March 2012

  • Afonine, Pavel V.; Grosse-Kunstleve, Ralf W.; Echols, Nathaniel
  • Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4
  • DOI: 10.1107/S0907444912001308

Web-based visualisation and analysis of 3D electron-microscopy data from EMDB and PDB
journal, November 2013

  • Lagerstedt, Ingvar; Moore, William J.; Patwardhan, Ardan
  • Journal of Structural Biology, Vol. 184, Issue 2
  • DOI: 10.1016/j.jsb.2013.09.021

A reciprocal-space method for calculating a molecular envelope using the algorithm of B.C. Wang
journal, January 1987

  • Leslie, A. G. W.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 43, Issue 1
  • DOI: 10.1107/S0108767387099720

Improved methods for building protein models in electron density maps and the location of errors in these models
journal, March 1991

  • Jones, T. A.; Zou, J. Y.; Cowan, S. W.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 47, Issue 2, p. 110-119
  • DOI: 10.1107/S0108767390010224

Likelihood-based refinement. I. Irremovable model errors
journal, May 2002

  • Lunin, V. Y.; Afonine, P. V.; Urzhumtsev, A. G.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 58, Issue 3
  • DOI: 10.1107/S0108767302001046

Automated map sharpening by maximization of detail and connectivity
journal, May 2018

  • Terwilliger, Thomas C.; Sobolev, Oleg V.; Afonine, Pavel V.
  • Acta Crystallographica Section D Structural Biology, Vol. 74, Issue 6
  • DOI: 10.1107/S2059798318004655

Neutron scattering lengths and cross sections
journal, January 1992


Outcome of the First Electron Microscopy Validation Task Force Meeting
journal, February 2012


Statistical quality indicators for electron-density maps
journal, March 2012

  • Tickle, Ian J.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4
  • DOI: 10.1107/S0907444911035918

Tricubic interpolation in three dimensions
journal, January 2005

  • Lekien, F.; Marsden, J.
  • International Journal for Numerical Methods in Engineering, Vol. 63, Issue 3
  • DOI: 10.1002/nme.1296

4.4 Å Resolution Cryo-EM structure of human mTOR Complex 1
journal, December 2016


Cryo-EM structure of the large subunit of the spinach chloroplast ribosome
journal, October 2016

  • Ahmed, Tofayel; Yin, Zhan; Bhushan, Shashi
  • Scientific Reports, Vol. 6, Issue 1
  • DOI: 10.1038/srep35793

Resolution and Probabilistic Models of Components in CryoEM Maps of Mature P22 Bacteriophage
journal, February 2016

  • Pintilie, Grigore; Chen, Dong-Hua; Haase-Pettingell, Cameron A.
  • Biophysical Journal, Vol. 110, Issue 4
  • DOI: 10.1016/j.bpj.2015.11.3522

2.8-Å Cryo-EM Structure of the Large Ribosomal Subunit from the Eukaryotic Parasite Leishmania
journal, July 2016


Structure of the E. coli ribosome–EF-Tu complex at <3 Å resolution by Cs-corrected cryo-EM
journal, February 2015

  • Fischer, Niels; Neumann, Piotr; Konevega, Andrey L.
  • Nature, Vol. 520, Issue 7548
  • DOI: 10.1038/nature14275

Electron Scattering Factors of Ions and their Parameterization
journal, July 1998

  • Peng, L. -M.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 54, Issue 4
  • DOI: 10.1107/S0108767398001901

Improved Fourier coefficients for maps using phases from partial structures with errors
journal, May 1986

  • Read, R. J.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 42, Issue 3
  • DOI: 10.1107/S0108767386099622

Restrained real-space macromolecular atomic refinement using a new resolution-dependent electron-density function
journal, January 1995

  • Chapman, M. S.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 51, Issue 1
  • DOI: 10.1107/S0108767394007130

Fitting atomic models into electron-microscopy maps
journal, October 2000

  • Rossmann, Michael G.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 56, Issue 10
  • DOI: 10.1107/S0907444900009562

3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM
journal, August 2016

  • Chua, Eugene Y. D.; Vogirala, Vinod K.; Inian, Oviya
  • Nucleic Acids Research, Vol. 44, Issue 17
  • DOI: 10.1093/nar/gkw708

The protein data bank: A computer-based archival file for macromolecular structures
journal, May 1977

  • Bernstein, Frances C.; Koetzle, Thomas F.; Williams, Graheme J. B.
  • Journal of Molecular Biology, Vol. 112, Issue 3
  • DOI: 10.1016/S0022-2836(77)80200-3

Bounding a molecule in a noisy synthesis
journal, January 1989

  • Urzhumtsev, A. G.; Lunin, V. Yu.; Luzyanina, T. B.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 45, Issue 1
  • DOI: 10.1107/S0108767388008955

Use of knowledge-based restraints in phenix.refine to improve macromolecular refinement at low resolution
journal, March 2012

  • Headd, Jeffrey J.; Echols, Nathaniel; Afonine, Pavel V.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4
  • DOI: 10.1107/S0907444911047834

Metrics for comparison of crystallographic maps
journal, September 2014

  • Urzhumtsev, Alexandre; Afonine, Pavel V.; Lunin, Vladimir Y.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 70, Issue 10
  • DOI: 10.1107/S1399004714016289

New analytical scattering-factor functions for free atoms and ions
journal, May 1995

  • Waasmaier, D.; Kirfel, A.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 51, Issue 3
  • DOI: 10.1107/S0108767394013292

Interpretation of ensembles created by multiple iterative rebuilding of macromolecular models
journal, April 2007

  • Terwilliger, Thomas C.; Grosse-Kunstleve, Ralf W.; Afonine, Pavel V.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 63, Issue 5
  • DOI: 10.1107/S0907444907009791

Structural Plasticity of the Protein Plug That Traps Newly Packaged Genomes in Podoviridae Virions
journal, November 2015

  • Bhardwaj, Anshul; Sankhala, Rajeshwer S.; Olia, Adam S.
  • Journal of Biological Chemistry, Vol. 291, Issue 1
  • DOI: 10.1074/jbc.M115.696260

Portal protein functions akin to a DNA-sensor that couples genome-packaging to icosahedral capsid maturation
journal, January 2017

  • Lokareddy, Ravi K.; Sankhala, Rajeshwer S.; Roy, Ankoor
  • Nature Communications, Vol. 8, Issue 1
  • DOI: 10.1038/ncomms14310

Macromolecular crystallographic estructure refinement [Macromolecular crystallographic estructure refinement]
journal, April 2015


Tools for the cryo-EM gold rush: going from the cryo-EM map to the atomistic model
journal, December 2017

  • Kim, Doo Nam; Sanbonmatsu, Karissa Y.
  • Bioscience Reports, Vol. 37, Issue 6
  • DOI: 10.1042/BSR20170072

Structure of the STRA6 receptor for retinol uptake
journal, August 2016


A real-space refinement procedure for proteins
journal, September 1971


MolProbity : all-atom structure validation for macromolecular crystallography
journal, December 2009

  • Chen, Vincent B.; Arendall, W. Bryan; Headd, Jeffrey J.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 1
  • DOI: 10.1107/S0907444909042073

Coot model-building tools for molecular graphics
journal, November 2004

  • Emsley, Paul; Cowtan, Kevin
  • Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12, p. 2126-2132
  • DOI: 10.1107/S0907444904019158

Crystallographic R Factor Refinement by Molecular Dynamics
journal, January 1987


Introduction to macromolecular refinement
journal, November 2004

  • Tronrud, Dale. E.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12
  • DOI: 10.1107/S090744490402356X

Model morphing and sequence assignment after molecular replacement
journal, October 2013

  • Terwilliger, Thomas C.; Read, Randy J.; Adams, Paul D.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 69, Issue 11
  • DOI: 10.1107/S0907444913017770

Features and development of Coot
journal, March 2010

  • Emsley, P.; Lohkamp, B.; Scott, W. G.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4
  • DOI: 10.1107/S0907444910007493

EMRinger: side chain–directed model and map validation for 3D cryo-electron microscopy
journal, August 2015

  • Barad, Benjamin A.; Echols, Nathaniel; Wang, Ray Yu-Ruei
  • Nature Methods, Vol. 12, Issue 10
  • DOI: 10.1038/nmeth.3541

Atomic model of a cypovirus built from cryo-EM structure provides insight into the mechanism of mRNA capping
journal, January 2011

  • Cheng, L.; Sun, J.; Zhang, K.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 4
  • DOI: 10.1073/pnas.1014995108

Programming new geometry restraints: parallelity of atomic groups
journal, July 2015

  • Sobolev, Oleg V.; Afonine, Pavel V.; Adams, Paul D.
  • Journal of Applied Crystallography, Vol. 48, Issue 4
  • DOI: 10.1107/S1600576715010432

MAIN software for density averaging, model building, structure refinement and validation
journal, June 2013


UCSF ChimeraX: Meeting modern challenges in visualization and analysis: UCSF ChimeraX Visualization System
journal, September 2017

  • Goddard, Thomas D.; Huang, Conrad C.; Meng, Elaine C.
  • Protein Science, Vol. 27, Issue 1
  • DOI: 10.1002/pro.3235

PHENIX: a comprehensive Python-based system for macromolecular structure solution
journal, January 2010

  • Adams, Paul D.; Afonine, Pavel V.; Bunkóczi, Gábor
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 213-221
  • DOI: 10.1107/S0907444909052925

A graphics model building and refinement system for macromolecules
journal, August 1978


Combining Electron Microscopic with X-Ray Crystallographic Structures
journal, December 2001

  • Rossmann, Michael G.; Bernal, Ricardo; Pletnev, Sergei V.
  • Journal of Structural Biology, Vol. 136, Issue 3
  • DOI: 10.1006/jsbi.2002.4435

Validated near-atomic resolution structure of bacteriophage epsilon15 derived from cryo-EM and modeling
journal, July 2013

  • Baker, M. L.; Hryc, C. F.; Zhang, Q.
  • Proceedings of the National Academy of Sciences, Vol. 110, Issue 30
  • DOI: 10.1073/pnas.1309947110

Improvement of protein phases by coarse model modification
journal, May 1984

  • Lunin, V. Yu.; Urzhumtsev, A. G.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 40, Issue 3
  • DOI: 10.1107/S0108767384000544

Fast and accurate conversion of atomic models into electron density maps
journal, January 2015

  • O. S. Sorzano, Carlos; Vargas, Javier; Otón, Joaquín
  • AIMS Biophysics, Vol. 2, Issue 1
  • DOI: 10.3934/biophy.2015.1.8

`Atomic resolution': a badly abused term in structural biology
journal, March 2017

  • Wlodawer, Alexander; Dauter, Zbigniew
  • Acta Crystallographica Section D Structural Biology, Vol. 73, Issue 4
  • DOI: 10.1107/S205979831700225X

Validation of Structures in the Protein Data Bank
journal, December 2017


The use of molecular-replacement phases for the refinement of the human rhinovirus 14 structure
journal, May 1988

  • Arnold, E.; Rossmann, M. G.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 44, Issue 3
  • DOI: 10.1107/S0108767387011875

The Computational Crystallography Toolbox : crystallographic algorithms in a reusable software framework
journal, January 2002

  • Grosse-Kunstleve, Ralf W.; Sauter, Nicholas K.; Moriarty, Nigel W.
  • Journal of Applied Crystallography, Vol. 35, Issue 1
  • DOI: 10.1107/S0021889801017824

A number of real-space torsion-angle refinement techniques for proteins, nucleic acids, ligands and solvent
journal, January 2001

  • Oldfield, Thomas J.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 57, Issue 1
  • DOI: 10.1107/S0907444900014098

Structure of the TRPV1 ion channel determined by electron cryo-microscopy
journal, December 2013


Automatic multiple-zone rigid-body refinement with a large convergence radius
journal, July 2009

  • Afonine, Pavel V.; Grosse-Kunstleve, Ralf W.; Urzhumtsev, Alexandre
  • Journal of Applied Crystallography, Vol. 42, Issue 4
  • DOI: 10.1107/S0021889809023528

Tools for macromolecular model building and refinement into electron cryo-microscopy reconstructions
journal, January 2015

  • Brown, Alan; Long, Fei; Nicholls, Robert A.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 71, Issue 1
  • DOI: 10.1107/S1399004714021683

Model bias in macromolecular crystal structures
journal, November 1992

  • Hodel, A.; Kim, S. H.; Brünger, A. T.
  • Acta Crystallographica Section A Foundations of Crystallography, Vol. 48, Issue 6
  • DOI: 10.1107/S0108767392006044

KING (Kinemage, Next Generation): A versatile interactive molecular and scientific visualization program
journal, November 2009

  • Chen, Vincent B.; Davis, Ian W.; Richardson, David C.
  • Protein Science, Vol. 18, Issue 11
  • DOI: 10.1002/pro.250

The Protein Data Bank
journal, January 2000


Free R value: a novel statistical quantity for assessing the accuracy of crystal structures
journal, January 1992


Between objectivity and subjectivity
journal, February 1990

  • Bränd´en, Carl-Ivar; Alwyn Jones, T.
  • Nature, Vol. 343, Issue 6260
  • DOI: 10.1038/343687a0

Automated Protein–Ligand Crystallography for Structure-Based Drug Design
journal, August 2006

  • Mooij, Wijnand T. M.; Hartshorn, Michael J.; Tickle, Ian J.
  • ChemMedChem, Vol. 1, Issue 8
  • DOI: 10.1002/cmdc.200600074