X-ray structure determination using low-resolution electron microscopy maps for molecular replacement

Jackson, Ryan N.; McCoy, Airlie J.; Terwilliger, Thomas C.; Read, Randy J.; Wiedenheft, Blake

doi:10.1038/nprot.2015.069

Title: X-ray structure determination using low-resolution electron microscopy maps for molecular replacement

Journal Article · Thu Jul 30 00:00:00 EDT 2015 · Nature Protocols

DOI:https://doi.org/10.1038/nprot.2015.069· OSTI ID:1254834

Jackson, Ryan N. ^[1]; McCoy, Airlie J. ^[2];

^[3]; Read, Randy J. ^[2]; Wiedenheft, Blake ^[1]

Montana State Univ., Bozeman, MT (United States)
Univ. of Cambridge (United Kingdom)
Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

Structures of multi-subunit macromolecular machines are primarily determined by either electron microscopy (EM) or X-ray crystallography. In many cases, a structure for a complex can be obtained at low resolution (at a coarse level of detail) with EM and at higher resolution (with finer detail) by X-ray crystallography. The integration of these two structural techniques is becoming increasingly important for generating atomic models of macromolecular complexes. A low-resolution EM image can be a powerful tool for obtaining the "phase" information that is missing from an X-ray crystallography experiment, however integration of EM and X-ray diffraction data has been technically challenging. Here we show a step-by-step protocol that explains how low-resolution EM maps can be placed in the crystallographic unit cell by molecular replacement, and how initial phases computed from the placed EM density are extended to high resolution by averaging maps over non-crystallographic symmetry. As the resolution gap between EM and Xray crystallography continues to narrow, the use of EM maps to help with X-ray crystal structure determination, as described in this protocol, will become increasingly effective.

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Research Organization:: Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)

Sponsoring Organization:: USDOE

Grant/Contract Number:: AC52-06NA25396; GM110732; GM108888; EPS-110134

OSTI ID:: 1254834

Report Number(s):: LA-UR-14-28194; nprot.2015.069

Journal Information:: Nature Protocols, Vol. 10, Issue 9; ISSN 1754-2189

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 17 works

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Immunogenicity and structures of a rationally designed prefusion MERS-CoV spike antigen Pallesen, Jesper; Wang, Nianshuang; Corbett, Kizzmekia S. Proceedings of the National Academy of Sciences, Vol. 114, Issue 35 https://doi.org/10.1073/pnas.1707304114	journal	August 2017
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