RT Journal Article
SR Electronic
T1 Analysis of Local Variability and Allostery in Macromolecular Assemblies using Cryo-EM and Focused Classification
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 365940
DO 10.1101/365940
A1 Zhang, Cheng
A1 Cantara, William
A1 Jeon, Youngmin
A1 Musier-Forsyth, Karin
A1 Grigorieff, Nikolaus
A1 Lyumkis, Dmitry
YR 2018
UL http://biorxiv.org/content/early/2018/07/28/365940.abstract
AB Single-particle electron cryo-microscopy and computational image classification can be used to analyze structural variability in macromolecules and their assemblies. In some cases, a particle may contain different regions that each display a range of distinct conformations. We have developed strategies, implemented within the Frealign and cisTEM image processing packages, to focus classify on specific regions of a particle and detect potential covariance. The strategies are based on masking the region of interest using either a 2-D mask applied to reference projections and particle images, or a 3-D mask applied to the 3-D volume. We show that focused classification approaches can be used to study structural allostery, a concept that is likely to gain more importance as datasets grow in size, allowing the distinction of more structural states and smaller differences between states. Finally, we apply the approaches to an experimental dataset containing the HIV-1 Transactivation Response (TAR) element RNA fused into the large bacterial ribosomal subunit, to deconvolve structural mobility within localized regions of interest.HighlightsDescription of different image classification strategies in single-particle cryo-EMQuantitative evaluation of two classification methods using simulated dataApplication of the two classification methods to an experimental dataset