6PKP

MicroED structure of proteinase K from a platinum-coated, polished, single lamella at 1.91A resolution (#10)

Summary for 6PKP

• Entry DOI: 10.2210/pdb6pkp/pdb
• EMDB information: 20356 20357 20358 20359 20360 20361 20362 20363 20364 20365 20366
• Descriptor: Proteinase K (2 entities in total)
• Functional Keywords: hydrolase
• Biological source: Parengyodontium album (Engyodontium album, Tritirachium album)
• Total number of polymer chains: 1
• Total formula weight: 28930.78

Authors

Martynowycz, M.W.,Zhao, W.,Hattne, J.,Jensen, G.J.,Gonen, T. (deposition date: 2019-06-29, release date: 2019-09-04, Last modification date: 2019-12-18)

Primary citation

Martynowycz, M.W.,Zhao, W.,Hattne, J.,Jensen, G.J.,Gonen, T.
Qualitative Analyses of Polishing and Precoating FIB Milled Crystals for MicroED.
Structure, 27:1594-, 2019

PubMed Abstract: Microcrystal electron diffraction (MicroED) leverages the strong interaction between matter and electrons to determine protein structures from vanishingly small crystals. This strong interaction limits the thickness of crystals that can be investigated by MicroED, mainly due to absorption. Recent studies have demonstrated that focused ion-beam (FIB) milling can thin crystals into ideal-sized lamellae; however, it is not clear how to best apply FIB milling for MicroED. Here, the effects of polishing the lamellae, whereby the last few nanometers are milled away using a low-current gallium beam, are explored in both the platinum-precoated and uncoated samples. Our results suggest that precoating samples with a thin layer of platinum followed by polishing the crystal surfaces prior to data collection consistently led to superior results as indicated by higher signal-to-noise ratio, higher resolution, and better refinement statistics. This study lays the foundation for routine and reproducible methodology for sample preparation in MicroED.

PubMed: 31422911
DOI: 10.1016/j.str.2019.07.004

Experimental method

ELECTRON CRYSTALLOGRAPHY (1.91 Å)