Protein structural ensembles are revealed by redefining X-ray electron density noise

Proc Natl Acad Sci U S A. 2014 Jan 7;111(1):237-42. doi: 10.1073/pnas.1302823110. Epub 2013 Dec 20.


To increase the power of X-ray crystallography to determine not only the structures but also the motions of biomolecules, we developed methods to address two classic crystallographic problems: putting electron density maps on the absolute scale of e(-)/Å(3) and calculating the noise at every point in the map. We find that noise varies with position and is often six to eight times lower than thresholds currently used in model building. Analyzing the rescaled electron density maps from 485 representative proteins revealed unmodeled conformations above the estimated noise for 45% of side chains and a previously hidden, low-occupancy inhibitor of HIV capsid protein. Comparing the electron density maps in the free and nucleotide-bound structures of three human protein kinases suggested that substrate binding perturbs distinct intrinsic allosteric networks that link the active site to surfaces that recognize regulatory proteins. These results illustrate general approaches to identify and analyze alternative conformations, low-occupancy small molecules, solvent distributions, communication pathways, and protein motions.

Keywords: Ringer; electron number density; molecular motions; protein dynamics; refinement against perturbed input data.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Allosteric Site
  • Anisotropy
  • Bacteriophage T4 / chemistry
  • Casein Kinase II / chemistry
  • Catalytic Domain
  • Computer Simulation
  • Crystallography, X-Ray / methods*
  • Cyclin-Dependent Kinase 2 / chemistry
  • Death-Associated Protein Kinases / chemistry
  • Electrons*
  • Humans
  • Image Processing, Computer-Assisted / methods
  • Interleukin-1beta / chemistry
  • Motion
  • Muramidase / chemistry
  • Protein Conformation
  • Protein Kinases / chemistry*
  • Proteins / chemistry*
  • Receptor Protein-Tyrosine Kinases / chemistry
  • Receptor, EphA3
  • Reproducibility of Results
  • Scorpion Venoms / chemistry
  • gag Gene Products, Human Immunodeficiency Virus / chemistry


  • Interleukin-1beta
  • Proteins
  • Scorpion Venoms
  • gag Gene Products, Human Immunodeficiency Virus
  • Protein Kinases
  • EPHA3 protein, human
  • Receptor Protein-Tyrosine Kinases
  • Receptor, EphA3
  • CSNK2A1 protein, human
  • Casein Kinase II
  • DAPK1 protein, human
  • Death-Associated Protein Kinases
  • CDK2 protein, human
  • Cyclin-Dependent Kinase 2
  • Muramidase