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PDBsum entry 2uuf

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protein ligands metals Protein-protein interface(s) links
Hydrolase/hydrolase inhibitor PDB id
2uuf
Jmol
Contents
Protein chains
28 a.a. *
252 a.a. *
Ligands
ASP-PHE-GLU-GLU-
ILE-PRO-GLU-GLU-
TYS-LEU
Metals
_CA
_NA
Waters ×346
* Residue conservation analysis
PDB id:
2uuf
Name: Hydrolase/hydrolase inhibitor
Title: Thrombin-hirugen binary complex at 1.26a resolution
Structure: Human alpha thrombin. Chain: a. Fragment: residues 328-363. Thrombin. Chain: b. Fragment: residues 364-622. Synonym: human alpha thrombin, coagulation factor ii. Hirudin i. Chain: h.
Source: Homo sapiens. Human. Organism_taxid: 9606. Tissue: blood plasma. Synthetic: yes. Hirudo medicinalis. Medicinal leech. Organism_taxid: 6421. Other_details: this polypeptide chain was chemically synthe
Resolution:
1.26Å     R-factor:   0.146     R-free:   0.194
Authors: H.U.Ahmed,M.P.Blakeley,M.Cianci,D.W.J.Cruickshank,J.A.Hubbar J.R.Helliwell
Key ref:
H.U.Ahmed et al. (2007). The determination of protonation states in proteins. Acta Crystallogr D Biol Crystallogr, 63, 906-922. PubMed id: 17642517 DOI: 10.1107/S0907444907029976
Date:
02-Mar-07     Release date:   04-Sep-07    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P00734  (THRB_HUMAN) -  Prothrombin
Seq:
Struc:
 
Seq:
Struc:
622 a.a.
28 a.a.
Protein chain
Pfam   ArchSchema ?
P00734  (THRB_HUMAN) -  Prothrombin
Seq:
Struc:
 
Seq:
Struc:
622 a.a.
252 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: Chains A, B: E.C.3.4.21.5  - Thrombin.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Preferential cleavage: Arg-|-Gly; activates fibrinogen to fibrin and releases fibrinopeptide A and B.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     extracellular region   1 term 
  Biological process     blood coagulation   2 terms 
  Biochemical function     catalytic activity     3 terms  

 

 
DOI no: 10.1107/S0907444907029976 Acta Crystallogr D Biol Crystallogr 63:906-922 (2007)
PubMed id: 17642517  
 
 
The determination of protonation states in proteins.
H.U.Ahmed, M.P.Blakeley, M.Cianci, D.W.Cruickshank, J.A.Hubbard, J.R.Helliwell.
 
  ABSTRACT  
 
The protonation states of aspartic acids and glutamic acids as well as histidine are investigated in four X-ray cases: Ni,Ca concanavalin A at 0.94 A, a thrombin-hirugen binary complex at 1.26 A resolution and two thrombin-hirugen-inhibitor ternary complexes at 1.32 and 1.39 A resolution. The truncation of the Ni,Ca concanavalin A data at various test resolutions between 0.94 and 1.50 A provided a test comparator for the ;unknown' thrombin-hirugen carboxylate bond lengths. The protonation states of aspartic acids and glutamic acids can be determined (on the basis of convincing evidence) even to the modest resolution of 1.20 A as exemplified by our X-ray crystal structure refinements of Ni and Mn concanavalin A and also as indicated in the 1.26 A structure of thrombin, both of which are reported here. The protonation-state indication of an Asp or a Glu is valid provided that the following criteria are met (in order of importance). (i) The acidic residue must have a single occupancy. (ii) Anisotropic refinement at a minimum diffraction resolution of 1.20 A (X-ray data-to-parameter ratio of approximately 3.5:1) is required. (iii) Both of the bond lengths must agree with the expectation (i.e. dictionary values), thus allowing some relaxation of the bond-distance standard uncertainties required to approximately 0.025 A for a '3sigma' determination or approximately 0.04 A for a '2sigma' determination, although some variation of the expected bond-distance values must be allowed according to the microenvironment of the hydrogen of interest. (iv) Although the F(o) - F(c) map peaks are most likely to be unreliable at the resolution range around 1.20 A, if admitted as evidence the peak at the hydrogen position must be greater than or equal to 2.5 sigma and in the correct geometry. (v) The atomic B factors need to be less than 10 A(2) for bond-length differentiation; furthermore, the C=O bond can also be expected to be observed with continuous 2F(o) - F(c) electron density and the C-OH bond with discontinuous electron density provided that the atomic B factors are less than approximately 20 A(2) and the contour level is increased. The final decisive option is to carry out more than one experiment, e.g. multiple X-ray crystallography experiments and ideally neutron crystallography. The complementary technique of neutron protein crystallography has provided evidence of the protonation states of histidine and acidic residues in concanavalin A and also the correct orientations of asparagine and glutamine side chains. Again, the truncation of the neutron data at various test resolutions between 2.5 and 3.0 A, even 3.25 and 3.75 A resolution, examines the limits of the neutron probe. These various studies indicate a widening of the scope of both X-ray and neutron probes in certain circumstances to elucidate the protonation states in proteins.
 
  Selected figure(s)  
 
Figure 6.
Figure 6 Asp28 carboxyl-group bond lengths (restrained) versus resolution for Ni,Ca concanavalin A and native (Mn,Ca) concanavalin A refined using SHELXL-97. As expected, with restraints on the dictionary 1.249 Å bond length dominates from 1.35 to 1.50 Å resolution. This is true in both cases, but Ni,Ca concanavalin A `behaves' better.
Figure 16.
Figure 16 Positive neutron F[o] - F[c] maps for His24 (all at 2.5 ) at various resolutions (calculated from the model with no D atoms included). (a) 2.2 Å resolution; (b) 2.5 Å resolution; (c) 2.75 Å resolution; (d) 3.0 Å resolution; (e) 3.25 Å resolution; (f) 3.5 Å resolution; (g) 3.75 Å resolution.
 
  The above figures are reprinted by permission from the IUCr: Acta Crystallogr D Biol Crystallogr (2007, 63, 906-922) copyright 2007.  
  Figures were selected by the author.  
 
 
    Author's comment    
 
  Synopsis The scope of X-ray and neutron protein crystallography to determine protonation states of functionally important amino acids is investigated at resolutions from 0.94Å to 1.50Å and from 2.2Å to 3.75Å respectively using concanavalin A as a test case. Key indicators are then combined to assign levels of confidence of such protonation states for three thrombin X-ray crystal structures at resolutions of 1.26Å, 1.32Å and 1.39Å. These various studies indicate a widening of the scope of both X-ray and neutron probes in certain circumstances to elucidate the protonation states in proteins.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20516624 L.Gabison, M.Chiadmi, M.El Hajji, B.Castro, N.Colloc'h, and T.Prangé (2010).
Near-atomic resolution structures of urate oxidase complexed with its substrate and analogues: the protonation state of the ligand.
  Acta Crystallogr D Biol Crystallogr, 66, 714-724.
PDB codes: 3l8w 3l9g 3lbg 3ld4
20606265 S.Noguchi (2010).
Isomerization mechanism of aspartate to isoaspartate implied by structures of Ustilago sphaerogena ribonuclease U2 complexed with adenosine 3'-monophosphate.
  Acta Crystallogr D Biol Crystallogr, 66, 843-849.
PDB codes: 3agn 3ago
19586953 E.Oksanen, M.P.Blakeley, F.Bonneté, M.T.Dauvergne, F.Dauvergne, and M.Budayova-Spano (2009).
Large crystal growth by thermal control allows combined X-ray and neutron crystallographic studies to elucidate the protonation states in Aspergillus flavus urate oxidase.
  J R Soc Interface, 6, S599-S610.  
19446529 G.D.Bailey, J.K.Hyun, A.K.Mitra, and R.L.Kingston (2009).
Proton-linked dimerization of a retroviral capsid protein initiates capsid assembly.
  Structure, 17, 737-748.
PDB codes: 3g0v 3g1g 3g1i 3g21 3g26 3g28 3g29
  19255472 R.M.Leal, S.C.Teixeira, M.P.Blakeley, E.P.Mitchell, and V.T.Forsyth (2009).
A preliminary neutron crystallographic study of an A-DNA crystal.
  Acta Crystallogr Sect F Struct Biol Cryst Commun, 65, 232-235.  
18656544 M.P.Blakeley, P.Langan, N.Niimura, and A.Podjarny (2008).
Neutron crystallography: opportunities, challenges, and limitations.
  Curr Opin Struct Biol, 18, 593-600.  
  18453706 S.C.Teixeira, M.P.Blakeley, R.M.Leal, E.P.Mitchell, and V.T.Forsyth (2008).
A preliminary neutron crystallographic study of thaumatin.
  Acta Crystallogr Sect F Struct Biol Cryst Commun, 64, 378-381.  
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