6PKK

MicroED structure of proteinase K from an uncoated, single lamella at 2.18A resolution (#5)


Experimental Data Snapshot

  • Method: ELECTRON CRYSTALLOGRAPHY
  • Resolution: 2.18 Å
  • R-Value Free: 0.265 
  • R-Value Work: 0.223 
  • R-Value Observed: 0.226 

wwPDB Validation   3D Report Full Report


This is version 1.2 of the entry. See complete history


Literature

Qualitative Analyses of Polishing and Precoating FIB Milled Crystals for MicroED.

Martynowycz, M.W.Zhao, W.Hattne, J.Jensen, G.J.Gonen, T.

(2019) Structure 27: 1594

  • DOI: https://doi.org/10.1016/j.str.2019.07.004
  • Primary Citation of Related Structures:  
    6PKJ, 6PKK, 6PKL, 6PKM, 6PKN, 6PKO, 6PKP, 6PKQ, 6PKR, 6PKS, 6PKT

  • 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.


  • Organizational Affiliation

    Howard Hughes Medical Institute, University of California Los Angeles, Los Angeles, CA, USA; Departments of Biological Chemistry and Physiology, University of California Los Angeles, Los Angeles, CA, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Proteinase K279Parengyodontium albumMutation(s): 0 
EC: 3.4.21.64
UniProt
Find proteins for P06873 (Parengyodontium album)
Explore P06873 
Go to UniProtKB:  P06873
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP06873
Sequence Annotations
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  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: ELECTRON CRYSTALLOGRAPHY
  • Resolution: 2.18 Å
  • R-Value Free: 0.265 
  • R-Value Work: 0.223 
  • R-Value Observed: 0.226 
  • Space Group: P 43 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 67.57α = 90
b = 67.57β = 90
c = 105.72γ = 90
Software Package:
Software NamePurpose
PHENIXrefinement
EM Software:
TaskSoftware PackageVersion
RECONSTRUCTIONPHENIX1.15.2
MODEL REFINEMENTPHENIX1.15

Structure Validation

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Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)United StatesR35 GM122588
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)United States2P50GM082545

Revision History  (Full details and data files)

  • Version 1.0: 2019-09-04
    Type: Initial release
  • Version 1.1: 2019-10-16
    Changes: Data collection, Database references
  • Version 1.2: 2019-12-18
    Changes: Author supporting evidence