PDBsum entry 1c5x

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Blood clotting PDB id
Jmol PyMol
Protein chain
248 a.a. *
FLC ×3
Waters ×1152
* Residue conservation analysis
PDB id:
Name: Blood clotting
Title: Structural basis for selectivity of a small molecule, s1- binding, sub-micromolar inhibitor of urokinase type plasminogen activator
Structure: Protein (urokinase-type plasminogen activator). Chain: a. Fragment: short chain. Engineered: yes. Protein (urokinase-type plasminogen activator). Chain: b. Fragment: catalytic domain. Engineered: yes
Source: Homo sapiens. Human. Organism_taxid: 9606. Expressed in: pichia pastoris. Expression_system_taxid: 4922.
Biol. unit: Dimer (from PQS)
1.75Å     R-factor:   0.195     R-free:   0.244
Authors: B.A.Katz,R.Mackman,C.Luong,K.Radika,A.Martelli, P.A.Sprengeler,J.Wang,H.Chan,L.Wong
Key ref:
B.A.Katz et al. (2000). Structural basis for selectivity of a small molecule, S1-binding, submicromolar inhibitor of urokinase-type plasminogen activator. Chem Biol, 7, 299-312. PubMed id: 10779411 DOI: 10.1016/S1074-5521(00)00104-6
22-Dec-99     Release date:   22-Dec-00    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P00749  (UROK_HUMAN) -  Urokinase-type plasminogen activator
431 a.a.
248 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.  - U-plasminogen activator.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Specific cleavage of Arg-|-Val bond in plasminogen to form plasmin.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     proteolysis   1 term 
  Biochemical function     serine-type endopeptidase activity     1 term  


DOI no: 10.1016/S1074-5521(00)00104-6 Chem Biol 7:299-312 (2000)
PubMed id: 10779411  
Structural basis for selectivity of a small molecule, S1-binding, submicromolar inhibitor of urokinase-type plasminogen activator.
B.A.Katz, R.Mackman, C.Luong, K.Radika, A.Martelli, P.A.Sprengeler, J.Wang, H.Chan, L.Wong.
BACKGROUND: Urokinase-type plasminogen activator (uPA) is a protease associated with tumor metastasis and invasion. Inhibitors of uPA may have potential as drugs for prostate, breast and other cancers. Therapeutically useful inhibitors must be selective for uPA and not appreciably inhibit the related, and structurally and functionally similar enzyme, tissue-type plasminogen activator (tPA), involved in the vital blood-clotting cascade. RESULTS: We produced mutagenically deglycosylated low molecular weight uPA and determined the crystal structure of its complex with 4-iodobenzo[b]thiophene 2-carboxamidine (K(i) = 0.21 +/- 0.02 microM). To probe the structural determinants of the affinity and selectivity of this inhibitor for uPA we also determined the structures of its trypsin and thrombin complexes, of apo-trypsin, apo-thrombin and apo-factor Xa, and of uPA, trypsin and thrombin bound by compounds that are less effective uPA inhibitors, benzo[b]thiophene-2-carboxamidine, thieno[2,3-b]-pyridine-2-carboxamidine and benzamidine. The K(i) values of each inhibitor toward uPA, tPA, trypsin, tryptase, thrombin and factor Xa were determined and compared. One selectivity determinant of the benzo[b]thiophene-2-carboxamidines for uPA involves a hydrogen bond at the S1 site to Ogamma(Ser190) that is absent in the Ala190 proteases, tPA, thrombin and factor Xa. Other subtle differences in the architecture of the S1 site also influence inhibitor affinity and enzyme-bound structure. CONCLUSIONS: Subtle structural differences in the S1 site of uPA compared with that of related proteases, which result in part from the presence of a serine residue at position 190, account for the selectivity of small thiophene-2-carboxamidines for uPA, and afford a framework for structure-based design of small, potent, selective uPA inhibitors.
  Selected figure(s)  
Figure 3.
Figure 3. (a) Structure of uPA–thieno[2,3-b]pyridine-2-carboxamidine, pH 6.5, at 1.65 Å resolution, on the (2|F[o]|–|F[c]|), α[c] map. The occupancies of conformation 1 (with green carbons) and 2 (light green carbons) are 66% and 34%, respectively. In the trypsin complex the corresponding occupancies are 54% and 46% at pH 5.5, and 31% and 69% at pH 8.2. (b) Superposition of uPA– and trypsin–thieno[2,3-b]pyridine-2-carboxamidine, pH 5.5. For clarity only the first conformation of the bound inhibitors is shown. In each complex the aromatic rings of the two bound conformations of the inhibitors lie in the same plane as one another (see Figure 4a). The two locations of the Hγ proton of Ser195 for the trypsin complex are shown.
Figure 4.
Figure 4. Structures and associated (2|F[o]|–|F[c]|), α[c] maps for (a) apo-trypsin, pH 7.7; and (b) apo-thrombin, pH 7.5, 1.47 Å resolution. In (b), the long water1–O[Trp215] distance (3.4 Å) is indicated in cyan.
  The above figures are reprinted by permission from Cell Press: Chem Biol (2000, 7, 299-312) copyright 2000.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19816553 N.Okimoto, N.Futatsugi, H.Fuji, A.Suenaga, G.Morimoto, R.Yanai, Y.Ohno, T.Narumi, and M.Taiji (2009).
High-performance drug discovery: computational screening by combining docking and molecular dynamics simulations.
  PLoS Comput Biol, 5, e1000528.  
18427113 D.Jiao, P.A.Golubkov, T.A.Darden, and P.Ren (2008).
Calculation of protein-ligand binding free energy by using a polarizable potential.
  Proc Natl Acad Sci U S A, 105, 6290-6295.  
18680100 N.Singh, and J.M.Briggs (2008).
Molecular dynamics simulations of Factor Xa: insight into conformational transition of its binding subsites.
  Biopolymers, 89, 1104-1113.  
17275887 A.V.Iakhiaev, A.Nalian, K.Koenig, and S.Idell (2007).
Thrombin-thrombomodulin inhibits prourokinase-mediated pleural mesothelial cell-dependent fibrinolysis.
  Thromb Res, 120, 715-725.  
17623865 V.Ramensky, A.Sobol, N.Zaitseva, A.Rubinov, and V.Zosimov (2007).
A novel approach to local similarity of protein binding sites substantially improves computational drug design results.
  Proteins, 69, 349-357.  
16387853 A.Fernández, R.Scott, and R.S.Berry (2006).
Packing defects as selectivity switches for drug-based protein inhibitors.
  Proc Natl Acad Sci U S A, 103, 323-328.  
17024701 M.Zentgraf, J.Fokkens, and C.A.Sotriffer (2006).
Addressing protein flexibility and ligand selectivity by "in situ cross-docking".
  ChemMedChem, 1, 1355-1359.  
16338411 A.Fernández (2005).
Incomplete protein packing as a selectivity filter in drug design.
  Structure, 13, 1829-1836.  
19003060 E.Atkins, S.Zamora, B.J.Candia, A.Baca, and R.A.Orlando (2005).
Development of a Mammalian suspension culture for expression of active recombinant human urokinase-type plasminogen activator.
  Cytotechnology, 49, 25-37.  
16199530 J.Tang, C.L.Yu, S.R.Williams, E.Springman, D.Jeffery, P.A.Sprengeler, A.Estevez, J.Sampang, W.Shrader, J.Spencer, W.Young, M.McGrath, and B.A.Katz (2005).
Expression, crystallization, and three-dimensional structure of the catalytic domain of human plasma kallikrein.
  J Biol Chem, 280, 41077-41089.
PDB codes: 2anw 2any
15545266 L.Jin, P.Pandey, R.E.Babine, J.C.Gorga, K.J.Seidl, E.Gelfand, D.T.Weaver, S.S.Abdel-Meguid, and J.E.Strickler (2005).
Crystal structures of the FXIa catalytic domain in complex with ecotin mutants reveal substrate-like interactions.
  J Biol Chem, 280, 4704-4712.
PDB codes: 1xx9 1xxd 1xxf
12962630 J.R.Somoza, J.D.Ho, C.Luong, M.Ghate, P.A.Sprengeler, K.Mortara, W.D.Shrader, D.Sperandio, H.Chan, M.E.McGrath, and B.A.Katz (2003).
The structure of the extracellular region of human hepsin reveals a serine protease domain and a novel scavenger receptor cysteine-rich (SRCR) domain.
  Structure, 11, 1123-1131.
PDB code: 1p57
11731301 B.A.Katz, P.A.Sprengeler, C.Luong, E.Verner, K.Elrod, M.Kirtley, J.Janc, J.R.Spencer, J.G.Breitenbucher, H.Hui, D.McGee, D.Allen, A.Martelli, and R.L.Mackman (2001).
Engineering inhibitors highly selective for the S1 sites of Ser190 trypsin-like serine protease drug targets.
  Chem Biol, 8, 1107-1121.
PDB codes: 1gj4 1gj5 1gj6 1gj7 1gj8 1gj9 1gja 1gjb 1gjc 1gjd
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.