Retrieving high-resolution information from disordered 2D crystals by single-particle cryo-EM.
basic_science · Level V
Where this comes from
- Record sourced from PubMed, PMID 30979902.
- Also identified by DOI 10.1038/s41467-019-09661-5 and PMC identifier 6461647.
- Licence recorded as CC BY.
- The licence permits redistribution, so the abstract is shown in full and the full text is available from the publisher.
Abstract
Electron crystallography can reveal the structure of membrane proteins within 2D crystals under close-to-native conditions. High-resolution structural information can only be reached if crystals are perfectly flat and highly ordered. In practice, such crystals are difficult to obtain. Available image unbending algorithms correct for disorder, but only perform well on images of non-tilted, flat crystals, while out-of-plane distortions are not addressed. Here, we present an approach that employs single-particle refinement procedures to locally unbend crystals in 3D. With this method, density maps of the MloK1 potassium channel with a resolution of 4 Å were obtained from images of 2D crystals that do not diffract beyond 10 Å. Furthermore, 3D classification allowed multiple structures to be resolved, revealing a series of MloK1 conformations within a single 2D crystal. This conformational heterogeneity explains the poor diffraction observed and is related to channel function. The approach is implemented in the FOCUS package.