@article {49180, title = {Analysis of global and site-specific radiation damage in cryo-EM.}, journal = {Structure (London, England : 1993)}, volume = {26}, year = {2018}, month = {2018 Apr 18}, pages = {759-66}, abstract = {

Micro-crystal electron diffraction (MicroED) combines the efficiency of electron scattering with diffraction to allow structure determination from nano-sized crystalline samples in cryoelectron microscopy (cryo-EM). It has been used to solve structures of a diverse set of biomolecules and materials, in some cases to sub-atomic resolution. However, little is known about the damaging effects of the electron beam on samples during such measurements. We assess global and site-specific damage from electron radiation on nanocrystals of proteinase K and of a prion hepta-peptide and find that the dynamics of electron-induced damage follow well-established trends observed in X-ray crystallography. Metal ions are perturbed, disulfide bonds are broken, and acidic side chains are decarboxylated while the\ diffracted intensities decay exponentially with increasing exposure. A better understanding of radiation damage in MicroED improves our assessment and processing of all types of cryo-EM data.

}, issn = {1878-4186}, doi = {10.1016/j.str.2018.03.021}, author = {Hattne, Johan and Shi, Dan and Glynn, Calina and Zee, Chih-Te and Gallagher-Jones, Marcus and Martynowycz, Michael W and Rodriguez, Jose A and Gonen, Tamir} } @article {49006, title = {Sub-{\r a}ngstr{\"o}m cryo-EM structure of a prion protofibril reveals a polar clasp.}, journal = {Nature Structural \& Molecular Biology}, year = {2018}, month = {2018 Jan 15}, abstract = {

The atomic structure of the infectious, protease-resistant, β-sheet-rich and fibrillar mammalian prion remains unknown. Through the cryo-EM method MicroED, we reveal the sub-{\r a}ngstr{\"o}m-resolution structure of a protofibril formed by a wild-type segment from the β2-α2 loop of the bank vole prion protein. The structure of this protofibril reveals a stabilizing network of hydrogen bonds that link polar zippers within a sheet, producing motifs we have named {\textquoteright}polar clasps{\textquoteright}.

}, issn = {1545-9985}, doi = {10.1038/s41594-017-0018-0}, author = {Gallagher-Jones, Marcus and Glynn, Calina and Boyer, David R and Martynowycz, Michael W and Hernandez, Evelyn and Miao, Jennifer and Zee, Chih-Te and Novikova, Irina V and Goldschmidt, Lukasz and McFarlane, Heather T and Helguera, Gustavo F and Evans, James E and Sawaya, Michael R and Cascio, Duilio and Eisenberg, David S and Gonen, Tamir and Rodriguez, Jose A} } @article {48603, title = {MicroED structures from micrometer thick protein crystals.}, journal = {bioRxiv}, year = {2017}, month = {2017 Jun 20}, abstract = {

Theoretical calculations suggest that crystals exceeding 100 nm thickness are excluded by dynamical scattering from successful structure determination using microcrystal electron diffraction (MicroED). These calculations are at odds with experimental results where MicroED structures have been determined from significantly thicker crystals. Here we systematically evaluate the influence of thickness on the accuracy of MicroED intensities and the ability to determine structures from protein crystals one micrometer thick. To do so, we compare ab initio structures of a human prion protein segment determined from thin crystals to those determined from crystals up to one micrometer thick. We also compare molecular replacement solutions from crystals of varying thickness for a larger globular protein, proteinase K. Our results indicate that structures can be reliably determined from crystals at least an order of magnitude thicker than previously suggested by simulation, opening the possibility for an even broader range of MicroED experiments.

}, doi = {10.1101/152504}, author = {Martynowycz, Michael and Glynn, Calina and Miao, Jennifer and de la Cruz, Michael Jason and Hattne, Johan and Shi, Dan and Cascio, Duilio and Rodriguez, Jose and Gonen, Tamir} }