3SQH

Crystal structure of prethrombin-2 mutant S195A in the the open form


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.20 Å
  • R-Value Free: 0.244 
  • R-Value Work: 0.189 
  • R-Value Observed: 0.192 

wwPDB Validation   3D Report Full Report


This is version 1.2 of the entry. See complete history


Literature

Crystal structures of prethrombin-2 reveal alternative conformations under identical solution conditions and the mechanism of zymogen activation.

Pozzi, N.Chen, Z.Zapata, F.Pelc, L.A.Barranco-Medina, S.Di Cera, E.

(2011) Biochemistry 50: 10195-10202

  • DOI: https://doi.org/10.1021/bi2015019
  • Primary Citation of Related Structures:  
    3SQE, 3SQH

  • PubMed Abstract: 

    Prethrombin-2 is the immediate zymogen precursor of the clotting enzyme thrombin, which is generated upon cleavage at R15 and separation of the A chain and catalytic B chain. The X-ray structure of prethrombin-2 determined in the free form at 1.9 Å resolution shows the 215-217 segment collapsed into the active site and occluding 49% of the volume available for substrate binding. Remarkably, some of the crystals harvested from the same crystallization well, under identical solution conditions, diffract to 2.2 Å resolution in the same space group but produce a structure in which the 215-217 segment moves >5 Å and occludes 24% of the volume available for substrate binding. The two alternative conformations of prethrombin-2 have the side chain of W215 relocating >9 Å within the active site and are relevant to the allosteric E*-E equilibrium of the mature enzyme. Another unanticipated feature of prethrombin-2 bears on the mechanism of prothrombin activation. R15 is found buried within the protein in ionic interactions with E14e, D14l, and E18, thereby making its exposure to solvent necessary for proteolytic attack and conversion to thrombin. On the basis of this structural observation, we constructed the E14eA/D14lA/E18A triple mutant to reduce the level of electrostatic coupling with R15 and promote zymogen activation. The mutation causes prethrombin-2 to spontaneously convert to thrombin, without the need for the snake venom ecarin or the physiological prothrombinase complex.


  • Organizational Affiliation

    Department of Biochemistry and Molecular Biology, Saint Louis University School of Medicine, St. Louis, Missouri 63104, United States.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Thrombin light chain, heavy chainA [auth E]290Homo sapiensMutation(s): 1 
Gene Names: F2human cDNA
EC: 3.4.21.5
UniProt & NIH Common Fund Data Resources
Find proteins for P00734 (Homo sapiens)
Explore P00734 
Go to UniProtKB:  P00734
PHAROS:  P00734
GTEx:  ENSG00000180210 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP00734
Sequence Annotations
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  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.20 Å
  • R-Value Free: 0.244 
  • R-Value Work: 0.189 
  • R-Value Observed: 0.192 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 44.359α = 90
b = 60.043β = 96.47
c = 49.691γ = 90
Software Package:
Software NamePurpose
CrystalCleardata collection
MOLREPphasing
REFMACrefinement
HKL-2000data reduction
HKL-2000data scaling

Structure Validation

View Full Validation Report



Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2011-11-30
    Type: Initial release
  • Version 1.1: 2012-05-23
    Changes: Database references
  • Version 1.2: 2023-09-13
    Changes: Data collection, Database references, Refinement description