5B35

Serial Femtosecond Crystallography (SFX) of Ground State Bacteriorhodopsin Crystallized from Bicelles Determined Using 7-keV X-ray Free Electron Laser (XFEL) at SACLA


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.35 Å
  • R-Value Free: 0.220 
  • R-Value Work: 0.166 
  • R-Value Observed: 0.169 

wwPDB Validation   3D Report Full Report


Ligand Structure Quality Assessment 


This is version 1.5 of the entry. See complete history


Literature

Membrane protein structure determination by SAD, SIR, or SIRAS phasing in serial femtosecond crystallography using an iododetergent

Nakane, T.Hanashima, S.Suzuki, M.Saiki, H.Hayashi, T.Kakinouchi, K.Sugiyama, S.Kawatake, S.Matsuoka, S.Matsumori, N.Nango, E.Kobayashi, J.Shimamura, T.Kimura, K.Mori, C.Kunishima, N.Sugahara, M.Takakyu, Y.Inoue, S.Masuda, T.Hosaka, T.Tono, K.Joti, Y.Kameshima, T.Hatsui, T.Yabashi, M.Inoue, T.Nureki, O.Iwata, S.Murata, M.Mizohata, E.

(2016) Proc Natl Acad Sci U S A 113: 13039-13044

  • DOI: https://doi.org/10.1073/pnas.1602531113
  • Primary Citation of Related Structures:  
    5B34, 5B35

  • PubMed Abstract: 

    The 3D structure determination of biological macromolecules by X-ray crystallography suffers from a phase problem: to perform Fourier transformation to calculate real space density maps, both intensities and phases of structure factors are necessary; however, measured diffraction patterns give only intensities. Although serial femtosecond crystallography (SFX) using X-ray free electron lasers (XFELs) has been steadily developed since 2009, experimental phasing still remains challenging. Here, using 7.0-keV (1.771 Å) X-ray pulses from the SPring-8 Angstrom Compact Free Electron Laser (SACLA), iodine single-wavelength anomalous diffraction (SAD), single isomorphous replacement (SIR), and single isomorphous replacement with anomalous scattering (SIRAS) phasing were performed in an SFX regime for a model membrane protein bacteriorhodopsin (bR). The crystals grown in bicelles were derivatized with an iodine-labeled detergent heavy-atom additive 13a (HAD13a), which contains the magic triangle, I3C head group with three iodine atoms. The alkyl tail was essential for binding of the detergent to the surface of bR. Strong anomalous and isomorphous difference signals from HAD13a enabled successful phasing using reflections up to 2.1-Å resolution from only 3,000 and 4,000 indexed images from native and derivative crystals, respectively. When more images were merged, structure solution was possible with data truncated at 3.3-Å resolution, which is the lowest resolution among the reported cases of SFX phasing. Moreover, preliminary SFX experiment showed that HAD13a successfully derivatized the G protein-coupled A2a adenosine receptor crystallized in lipidic cubic phases. These results pave the way for de novo structure determination of membrane proteins, which often diffract poorly, even with the brightest XFEL beams.


  • Organizational Affiliation

    Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Bacteriorhodopsin249Halobacterium salinarumMutation(s): 0 
Membrane Entity: Yes 
UniProt
Find proteins for B0R5N9 (Halobacterium salinarum (strain ATCC 29341 / DSM 671 / R1))
Explore B0R5N9 
Go to UniProtKB:  B0R5N9
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupB0R5N9
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 8 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
4QM
Query on 4QM

Download Ideal Coordinates CCD File 
C [auth A](3R,5S,7R,8R,9S,10S,12S,13R,14S,17R)-10,13-dimethyl-17-[(2R)-pentan-2-yl]-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthrene-3,7,12-triol
C24 H42 O3
PFZUIDNKXWIWBG-YHEMGIGTSA-N
RET
Query on RET

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B [auth A]RETINAL
C20 H28 O
NCYCYZXNIZJOKI-OVSJKPMPSA-N
R16
Query on R16

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J [auth A],
R [auth A]
HEXADECANE
C16 H34
DCAYPVUWAIABOU-UHFFFAOYSA-N
D12
Query on D12

Download Ideal Coordinates CCD File 
F [auth A],
O [auth A],
P [auth A]
DODECANE
C12 H26
SNRUBQQJIBEYMU-UHFFFAOYSA-N
D10
Query on D10

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E [auth A],
G [auth A],
H [auth A],
I [auth A],
K [auth A]
DECANE
C10 H22
DIOQZVSQGTUSAI-UHFFFAOYSA-N
DD9
Query on DD9

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N [auth A],
Q [auth A]
nonane
C9 H20
BKIMMITUMNQMOS-UHFFFAOYSA-N
OCT
Query on OCT

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D [auth A]N-OCTANE
C8 H18
TVMXDCGIABBOFY-UHFFFAOYSA-N
HP6
Query on HP6

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L [auth A],
M [auth A]
HEPTANE
C7 H16
IMNFDUFMRHMDMM-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

Unit Cell:
Length ( Å )Angle ( ˚ )
a = 46.2α = 90
b = 103β = 90
c = 128.7γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
Cheetahdata extraction
CrystFELdata processing
SHELXphasing

Structure Validation

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Ligand Structure Quality Assessment 


Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
X-ray Free-Electron Laser Priority Strategy Program (MEXT)JapanSACLA-SFX Project
Japanese Society of Technology ERATOJapanMurata Lipid Active Structure Project

Revision History  (Full details and data files)

  • Version 1.0: 2016-11-09
    Type: Initial release
  • Version 1.1: 2016-11-16
    Changes: Database references
  • Version 1.2: 2016-11-30
    Changes: Database references
  • Version 1.3: 2018-01-24
    Changes: Data collection, Database references
  • Version 1.4: 2023-09-06
    Changes: Data collection, Database references
  • Version 1.5: 2023-11-08
    Changes: Refinement description