3J7H

Structure of beta-galactosidase at 3.2-A resolution obtained by cryo-electron microscopy

Summary for 3J7H

• Entry DOI: 10.2210/pdb3j7h/pdb
• EMDB information: 5995
• Descriptor: Beta-galactosidase, MAGNESIUM ION (2 entities in total)
• Functional Keywords: hydrolase enzyme, homo-tetramer, protein complex, atomic resolution cryo-electron microscopy, direct electron detectors, single-particle cryo-em, 3d reconstruction, hydrolase
• Biological source: Escherichia coli K-12
• Total number of polymer chains: 4
• Total formula weight: 466507.16

Authors

Bartesaghi, A.,Matthies, D.,Banerjee, S.,Merk, A.,Subramaniam, S. (deposition date: 2014-06-30, release date: 2014-07-30, Last modification date: 2024-02-21)

Primary citation

Bartesaghi, A.,Matthies, D.,Banerjee, S.,Merk, A.,Subramaniam, S.
Structure of beta-galactosidase at 3.2- angstrom resolution obtained by cryo-electron microscopy.
Proc.Natl.Acad.Sci.USA, 111:11709-11714, 2014

PubMed Abstract: 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 Mg(2+) 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(-)/Å(2). In summary, we establish that it is feasible to use cryo-EM to determine near-atomic resolution structures of protein complexes (<500 kDa) with low symmetry, and that the residue-specific radiation damage that occurs with increasing electron dose can be monitored by using dose fractionation tools available with direct electron detector technology.

PubMed: 25071206
DOI: 10.1073/pnas.1402809111

Experimental method

ELECTRON MICROSCOPY (3.2 Å)