Workshop 8: Microcrystal Electron Diffraction (MicroED)

Date/Time: Friday, July 29, 2022 @ 8:00 AM PT
Location: ROOM TBA

Portland Marriott Downtown, Portland, Oregon

Organizational Contact: Tamir Gonen, Jessica Bruhn

 

     

 

Description

The aim of this workshop is to provide hands on training for microcrystal electron diffraction (microED) data collection. The focus of the workshop with be the current state of data collection, data reduction and phasing. Model building and structure refinement will not be specifically covered. Data reduction will be demonstrated with freely available programs (DIALS, XDS, MOSFILM, POINTLESS, etc.).

The morning sessions will include classroom lectures on the practical aspects of carrying out a microED experiment. Specific topics will include grid preparation, focused ion beam milling of crystals, data collection strategies, and the current state of data processing.

The afternoon session will focus on hands-on demonstration of imaging microcrystals, identifying suitable crystals for data collection and setting up continuous rotation experiments (data collection). This demonstration will use the software EPU-D. Groups of 10 will be allowed to take turns remotely controlling an electron microscope. The other groups will be working on self-guided tutorials for data reduction. Instructors will be circulating through the room while members work on their tutorials.

Morning sessions with introductions on sample preparation, data collection, and data reduction

In the many scientific endeavors that are driven by chemistry, unambiguous identification of molecules is of paramount importance. Over the past 50 years, NMR and other powerful spectroscopic techniques have been developed to address this challenge. While almost all of these techniques rely on inference of connectivity, the unambiguous determination of a small molecule’s structure requires X-ray and/or neutron diffraction studies. It is now possible to use cryo-electron microscopy (cryoEM) methods to obtain electron diffraction data from microcrystals, greatly expanding the type of samples amenable to crystal structure determination. This has been demonstrated with small and macromolecule targets. From simple powders of small molecules, with minimal sample preparation, it is possible to collect high-quality microcrystal electron diffraction (microED) data from crystals ∼100 nm thick resulting in atomic resolution (<1 Å) crystal structures in minutes from small molecules. Similar techniques result in high resolution crystal structures of macromolecules.

The morning sessions will focus on the practical aspects of carrying out a microED experiment. Specific topics will include sample preparation, focused ion beam milling of crystals, data collection strategies, and the current state of data processing.

Afternoon sessions with simultaneous data collection and reduction tutorials.

Currently, most microED data is processed in standard X-ray software familiar to many crystallographers. Successful execution of data reduction with these programs can be difficult, even for seasoned X-ray diffraction veterans. Workshop attendees will follow self-guided tutorials on how to process a representative small molecule and macromolecular target. In 1-hour time slots, groups of 10 will be allowed to visit a central computer to remotely control an electron microscope (see the timeline in section 6). The microscope will be from the Thermo Fisher fleet, and will require the use of EPU-D, a vendor-specific software.

In order to maximize instructional time, participants will be issued USB drives that will boot into a virtual Linux environment with all the software needed for data reduction. Tutorials will be printed in workbooks with step-by-step instructions on how to go from electron diffraction patterns to a reduced reflections file (HKL or MTZ for small and macromolecules respectively).

 

Instructors

Tamir Gonen
Jessica Bruhn
Johan Hattne
Mike Martynowycz
Max Clabbers
Sara Weaver
 

Schedule 

 Start EndInstructor/Topic 
8:00 9:00 Boot into Linux environment
9:00 9:30 Introduction  
9:30 10:00 Sample Preparation Lecture 
10:00 10:30 Data Collection Overview (EPU-D)
10:30 11:00 Break
11:00 5:00 Self-Guided & Hands On Sessions