CaltechAUTHORS
  A Caltech Library Service

Collection of continuous rotation MicroED Data from Ion Beam Milled Crystals of Any Size

Martynowycz, Michael W. and Zhao, Wei and Hattne, Johan and Jensen, Grant J. and Gonen, Tamir (2019) Collection of continuous rotation MicroED Data from Ion Beam Milled Crystals of Any Size. Structure, 27 (3). pp. 545-548. ISSN 0969-2126. PMCID PMC6476546. https://resolver.caltech.edu/CaltechAUTHORS:20180927-114225333

[img] PDF - Accepted Version
See Usage Policy.

979Kb
[img] PDF - Submitted Version
Creative Commons Attribution Non-commercial No Derivatives.

734Kb

Use this Persistent URL to link to this item: https://resolver.caltech.edu/CaltechAUTHORS:20180927-114225333

Abstract

Microcrystal electron diffraction (MicroED) allows for macromolecular structure solution from nanocrystals. To create crystals of suitable size for MicroED data collection, sample preparation typically involves sonication or pipetting a slurry of crystals from a crystallization drop. The resultant crystal fragments are fragile and the quality of the data that can be obtained from them is sensitive to subsequent sample preparation for cryoelectron microscopy as interactions in the water-air interface can damage crystals during blotting. Here, we demonstrate the use of a focused ion beam to generate lamellae of macromolecular protein crystals for continuous rotation MicroED that are of ideal thickness, easy to locate, and require no blotting optimization. In this manner, crystals of nearly any size may be scooped and milled to desired dimensions prior to data collection, thus streamlining the methodology for sample preparation for MicroED.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1016/j.str.2018.12.003DOIArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6476546PubMed CentralArticle
https://doi.org/10.1101/425611DOIDiscussion Paper
ORCID:
AuthorORCID
Jensen, Grant J.0000-0003-1556-4864
Additional Information:© 2018 Elsevier Ltd. Received 5 October 2018, Revised 19 November 2018, Accepted 5 December 2018, Available online 17 January 2019. The Gonen lab is supported by funds from the Howard Hughes Medical Institute. Work done in the Jensen lab was supported by NIH grant R35 GM122588. We would like to thank Ikaasa Suri (UCLA) for the help with sample preparation. Author Contributions: T.G. and G.J.J. conceived the project. M.W.M. prepared the protein crystals. W.Z. and M.W.M. collected the FIB/SEM data and milled the crystals. M.W.M. collected the MicroED data. M.W.M. and J.H. processed the data. The manuscript was written by M.W.M. and T.G. with contributions from all authors. M.W.M., G.J., and T.G. designed the experiments. Figures were prepared by M.W.M. and T.G. The authors declare no conflict of interests.
Funders:
Funding AgencyGrant Number
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
NIHR35 GM122588
Subject Keywords:Electron cryo-microscopy (cryoEM); MicroED; FIB-SEM; electron diffraction; electron crystallography
Issue or Number:3
PubMed Central ID:PMC6476546
Record Number:CaltechAUTHORS:20180927-114225333
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20180927-114225333
Official Citation:Michael W. Martynowycz, Wei Zhao, Johan Hattne, Grant J. Jensen, Tamir Gonen, Collection of Continuous Rotation MicroED Data from Ion Beam-Milled Crystals of Any Size, Structure, Volume 27, Issue 3, 2019, Pages 545-548.e2, ISSN 0969-2126, https://doi.org/10.1016/j.str.2018.12.003. (http://www.sciencedirect.com/science/article/pii/S0969212618304593)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:90014
Collection:CaltechAUTHORS
Deposited By: George Porter
Deposited On:27 Sep 2018 20:25
Last Modified:03 Oct 2019 20:21

Repository Staff Only: item control page