scholarly journals Machining protein microcrystals for structure determination by electron diffraction

2018 ◽  
Vol 115 (38) ◽  
pp. 9569-9573 ◽  
Author(s):  
Helen M. E. Duyvesteyn ◽  
Abhay Kotecha ◽  
Helen M. Ginn ◽  
Corey W. Hecksel ◽  
Emma V. Beale ◽  
...  

We demonstrate that ion-beam milling of frozen, hydrated protein crystals to thin lamella preserves the crystal lattice to near-atomic resolution. This provides a vehicle for protein structure determination, bridging the crystal size gap between the nanometer scale of conventional electron diffraction and micron scale of synchrotron microfocus beamlines. The demonstration that atomic information can be retained suggests that milling could provide such detail on sections cut from vitrified cells.

2015 ◽  
Vol 12 (4) ◽  
pp. 335-338 ◽  
Author(s):  
Ray Yu-Ruei Wang ◽  
Mikhail Kudryashev ◽  
Xueming Li ◽  
Edward H Egelman ◽  
Marek Basler ◽  
...  

Author(s):  
Emma V. Beale ◽  
David G. Waterman ◽  
Corey Hecksel ◽  
Jason van Rooyen ◽  
James B. Gilchrist ◽  
...  

Author(s):  
Emma V. Beale ◽  
David G. Waterman ◽  
Corey Hecksel ◽  
Jason van Rooyen ◽  
James B. Gilchrist ◽  
...  

AbstractMicro-Electron Diffraction (MicroED) has recently emerged as a powerful method for the analysis of biological structures at atomic resolution. This technique has been largely limited to protein nanocrystals which grow either as needles or plates measuring only a few hundred nanometres in thickness. Furthermore, traditional microED data processing uses established X-ray crystallography software that is not optimised for handling compound effects that are unique to electron diffraction data. Here, we present an integrated workflow for microED, from sample preparation by cryo-focused ion beam milling, through data collection with a standard Ceta-D detector, to data processing using the DIALS software suite, thus enabling routine atomic structure determination of protein crystals of any size and shape using microED. We demonstrate the effectiveness of the workflow by determining the structure of proteinase K to 2.0 Å resolution and show the advantage of using protein crystal lamellae over nanocrystals.


Nature ◽  
2020 ◽  
Vol 587 (7832) ◽  
pp. 157-161 ◽  
Author(s):  
Ka Man Yip ◽  
Niels Fischer ◽  
Elham Paknia ◽  
Ashwin Chari ◽  
Holger Stark

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