PDBsum entry 3dhk

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protein ligands metals Protein-protein interface(s) links
Blood clotting/hydrolase inhibitor PDB id
Protein chains
27 a.a. *
250 a.a. *
_NA ×2
Waters ×126
* Residue conservation analysis
PDB id:
Name: Blood clotting/hydrolase inhibitor
Title: Bisphenylic thrombin inhibitors
Structure: Thrombin light chain. Chain: l. Synonym: coagulation factor ii. Thrombin heavy chain. Chain: h. Synonym: coagulation factor ii. Hirudin variant-1. Chain: i. Fragment: residues 54-64.
Source: Homo sapiens. Human. Organism_taxid: 9606. Tissue: blood plasma. Synthetic: yes. Other_details: synthetic fragment of hirudin from hirudo me
1.73Å     R-factor:   0.197     R-free:   0.295
Authors: B.Baum,A.Heine,G.Klebe
Key ref:
B.Baum et al. (2009). Think twice: understanding the high potency of bis(phenyl)methane inhibitors of thrombin. J Mol Biol, 391, 552-564. PubMed id: 19520086 DOI: 10.1016/j.jmb.2009.06.016
18-Jun-08     Release date:   19-May-09    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P00734  (THRB_HUMAN) -  Prothrombin
622 a.a.
27 a.a.
Protein chain
Pfam   ArchSchema ?
P00734  (THRB_HUMAN) -  Prothrombin
622 a.a.
250 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: Chains L, H: E.C.  - 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: 10.1016/j.jmb.2009.06.016 J Mol Biol 391:552-564 (2009)
PubMed id: 19520086  
Think twice: understanding the high potency of bis(phenyl)methane inhibitors of thrombin.
B.Baum, L.Muley, A.Heine, M.Smolinski, D.Hangauer, G.Klebe.
Successful design of potent and selective protein inhibitors, in terms of structure-based drug design, strongly relies on the correct understanding of the molecular features determining the ligand binding to the target protein. We present a case study of serine protease inhibitors with a bis(phenyl)methane moiety binding into the S3 pocket. These inhibitors bind with remarkable potency to the active site of thrombin, the blood coagulation factor IIa. A combination of X-ray crystallography and isothermal titration calorimetry provides conclusive insights into the driving forces responsible for the surprisingly high potency of these inhibitors. Analysis of six well-resolved crystal structures (resolution 1.58-2.25 A) along with the thermodynamic data allows an explanation of the tight binding of the bis(phenyl)methane inhibitors. Interestingly, the two phenyl rings contribute to binding affinity for very different reasons - a fact that can only be elucidated by a structure-based approach. The first phenyl moiety occupies the hydrophobic S3 pocket, resulting in a mainly entropic advantage of binding. This observation is based on the displacement of structural water molecules from the S3 pocket that are observed in complexes with inhibitors that do not bind in the S3 pocket. The same classic hydrophobic effect cannot explain the enhanced binding affinity resulting from the attachment of the second, more solvent-exposed phenyl ring. For the bis(phenyl)methane inhibitors, an observed adaptive rotation of a glutamate residue adjacent to the S3 binding pocket attracted our attention. The rotation of this glutamate into salt-bridging distance with a lysine moiety correlates with an enhanced enthalpic contribution to binding for these highly potent thrombin binders. This explanation for the magnitude of the attractive force is confirmed by data retrieved by a Relibase search of several thrombin-inhibitor complexes deposited in the Protein Data Bank exhibiting similar molecular features. Special attention was attributed to putative changes in the protonation states of the interaction partners. For this purpose, two analogous inhibitors differing mainly in their potential to change the protonation state of a hydrogen-bond donor functionality were compared. Buffer dependencies of the binding enthalpy associated with complex formation could be traced by isothermal titration calorimetry, which revealed, along with analysis of the crystal structures (resolution 1.60 and 1.75 A), that a virtually compensating proton interchange between enzyme, inhibitor and buffer is responsible for the observed buffer-independent thermodynamic signatures.
  Selected figure(s)  
Figure 1.
Fig. 1. Chemical formulas of inhibitors 1 to 6 discussed in this study. Inhibitory potency towards human thrombin is given as kinetic inhibition constant K[i] with the SD from averaging of at least three measurements. Relative differences of the thermodynamic properties between inhibitors, as indicated by the arrows, are given in kilojoules per mole.
Figure 2.
Fig. 2. Inhibitor 3 in complex with human thrombin blocked at the exosite by hirugen. (a) View of the active site, with solvent-accessible surface of thrombin's heavy chain in pale green, light chain in blue and hirugen in violet. Two refined sodium ions are shown as yellow spheres, water molecules as red spheres and the inhibitor skeleton in orange sticks; heteroatoms are color-coded. The F[o] − F[c] difference electron density for 3 is depicted in blue at 2.5σ. (b) Active-site residues are represented by green sticks; ligand 3 is shown in orange. The red sphere represents the water molecule being involved in a hydrogen-bonding network. Hydrogen bonds are indicated as broken lines in magenta.
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2009, 391, 552-564) copyright 2009.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21157775 R.J.Falconer, and B.M.Collins (2011).
Survey of the year 2009: applications of isothermal titration calorimetry.
  J Mol Recognit, 24, 1.  
20695475 R.Baron, P.Setny, and J.A.McCammon (2010).
Water in cavity-ligand recognition.
  J Am Chem Soc, 132, 12091-12097.  
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