5UVL

Serial Millisecond Crystallography of Membrane and Soluble Protein Micro-crystals using Synchrotron Radiation


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.65 Å
  • R-Value Free: 0.243 
  • R-Value Work: 0.225 
  • R-Value Observed: 0.228 

wwPDB Validation   3D Report Full Report


This is version 1.6 of the entry. See complete history


Literature

Serial millisecond crystallography of membrane and soluble protein microcrystals using synchrotron radiation.

Martin-Garcia, J.M.Conrad, C.E.Nelson, G.Stander, N.Zatsepin, N.A.Zook, J.Zhu, L.Geiger, J.Chun, E.Kissick, D.Hilgart, M.C.Ogata, C.Ishchenko, A.Nagaratnam, N.Roy-Chowdhury, S.Coe, J.Subramanian, G.Schaffer, A.James, D.Ketwala, G.Venugopalan, N.Xu, S.Corcoran, S.Ferguson, D.Weierstall, U.Spence, J.C.H.Cherezov, V.Fromme, P.Fischetti, R.F.Liu, W.

(2017) IUCrJ 4: 439-454

  • DOI: https://doi.org/10.1107/S205225251700570X
  • Primary Citation of Related Structures:  
    5UVI, 5UVJ, 5UVK, 5UVL

  • PubMed Abstract: 

    Crystal structure determination of biological macromolecules using the novel technique of serial femtosecond crystallography (SFX) is severely limited by the scarcity of X-ray free-electron laser (XFEL) sources. However, recent and future upgrades render microfocus beamlines at synchrotron-radiation sources suitable for room-temperature serial crystallography data collection also. Owing to the longer exposure times that are needed at synchrotrons, serial data collection is termed serial millisecond crystallography (SMX). As a result, the number of SMX experiments is growing rapidly, with a dozen experiments reported so far. Here, the first high-viscosity injector-based SMX experiments carried out at a US synchrotron source, the Advanced Photon Source (APS), are reported. Microcrystals (5-20 µm) of a wide variety of proteins, including lysozyme, thaumatin, phycocyanin, the human A 2A adenosine receptor (A 2A AR), the soluble fragment of the membrane lipoprotein Flpp3 and proteinase K, were screened. Crystals suspended in lipidic cubic phase (LCP) or a high-molecular-weight poly(ethylene oxide) (PEO; molecular weight 8 000 000) were delivered to the beam using a high-viscosity injector. In-house data-reduction (hit-finding) software developed at APS as well as the SFX data-reduction and analysis software suites Cheetah and CrystFEL enabled efficient on-site SMX data monitoring, reduction and processing. Complete data sets were collected for A 2A AR, phycocyanin, Flpp3, proteinase K and lysozyme, and the structures of A 2A AR, phycocyanin, proteinase K and lysozyme were determined at 3.2, 3.1, 2.65 and 2.05 Å resolution, respectively. The data demonstrate the feasibility of serial millisecond crystallography from 5-20 µm crystals using a high-viscosity injector at APS. The resolution of the crystal structures obtained in this study was dictated by the current flux density and crystal size, but upcoming developments in beamline optics and the planned APS-U upgrade will increase the intensity by two orders of magnitude. These developments will enable structure determination from smaller and/or weakly diffracting microcrystals.


  • Organizational Affiliation

    School of Molecular Sciences and Biodesign Center for Applied Structural Discovery, Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Proteinase K279Parengyodontium albumMutation(s): 1 
Gene Names: PROK
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: X-RAY DIFFRACTION
  • Resolution: 2.65 Å
  • R-Value Free: 0.243 
  • R-Value Work: 0.225 
  • R-Value Observed: 0.228 
  • Space Group: P 43 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 68.3α = 90
b = 68.3β = 90
c = 108.2γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
CrystFELdata reduction
CrystFELdata scaling
Cootmodel building
MOLREPphasing
Cheetahdata collection

Structure Validation

View Full Validation Report



Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
Flinn FoundationUnited States1991
STC Program of the National Science Foundation through BioXFELUnited States1231306
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)United StatesR01GM095583
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)United StatesR01GM108635
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)United StatesU54GM094618
National Institutes of Health/National Cancer Institute (NIH/NCI)United StatesY1-CO-1020
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)United StatesY1-GM-1104
Department of Energy (DOE, United States)United StatesDE-AC02-06CH1135

Revision History  (Full details and data files)

  • Version 1.0: 2017-05-24
    Type: Initial release
  • Version 1.1: 2017-07-12
    Changes: Database references
  • Version 1.2: 2017-09-13
    Changes: Author supporting evidence
  • Version 1.3: 2018-04-18
    Changes: Data collection, Database references
  • Version 1.4: 2019-12-04
    Changes: Author supporting evidence
  • Version 1.5: 2022-04-13
    Changes: Author supporting evidence, Database references
  • Version 1.6: 2023-10-04
    Changes: Data collection, Refinement description