Structure of β-galactosidase at 3.2-Å resolution obtained by cryo-electron microscopy

Alberto Bartesaghi, Doreen Matthies, Soojay Banerjee, Alan Merk, Sriram Subramaniam

Research output: Contribution to journalArticlepeer-review

122 Scopus citations


We report the solution structure of Escherichia coli β-galactosidase (~465 kDa), solved at ~3.2-Å resolution by using single-particle cryo-electron microscopy (cryo-EM). Densities for most side chains, including those of residues in the active site, and a catalytic Mg2+ ion can be discerned in the map obtained by cryo-EM. The atomic model derived from our cryo-EM analysis closely matches the 1.7-Å crystal structure with a global rmsd of ~0.66 Å. There are significant local differences throughout the protein, with clear evidence for conformational changes resulting from contact zones in the crystal lattice. Inspection of the map reveals that although densities for residues with positively charged and neutral side chains are well resolved, systematically weaker densities are observed for residues with negatively charged side chains. We show that the weaker densities for negatively charged residues arise from their greater sensitivity to radiation damage from electron irradiation as determined by comparison of density maps obtained by using electron doses ranging from 10 to 30 e-2. In summary, we establish that it is feasible to use cryo-EM to determine near-atomic resolution structures of protein complexes (

Original languageEnglish (US)
Pages (from-to)11709-11714
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number32
StatePublished - Aug 12 2014
Externally publishedYes


  • 3D reconstruction
  • CTF determination
  • Frame alignment
  • Single-particle EM
  • Structure refinement

ASJC Scopus subject areas

  • General


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