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MicroED structure of proteinase K from merging low-dose, platinum pre-coated lamellae at 1.85A resolution (LDPT)MicroED structure of proteinase K from merging low-dose, platinum pre-coated lamellae at 1.85A resolution (LDPT)
Structural highlights
FunctionPRTK_PARAQ Hydrolyzes keratin at aromatic and hydrophobic residues. Publication Abstract from PubMedMicrocrystal 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. Qualitative Analyses of Polishing and Precoating FIB Milled Crystals for MicroED.,Martynowycz MW, Zhao W, Hattne J, Jensen GJ, Gonen T Structure. 2019 Oct 1;27(10):1594-1600.e2. doi: 10.1016/j.str.2019.07.004. Epub, 2019 Aug 15. PMID:31422911[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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