PDBsum entry 1rtf

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Serine protease PDB id
Protein chain
244 a.a. *
Waters ×168
* Residue conservation analysis
PDB id:
Name: Serine protease
Title: Complex of benzamidine with the catalytic domain of human two chain tissue plasminogen activator [(tc)-t-pa]
Structure: Two chain tissue plasminogen activator. Chain: a. Fragment: catalytic domain. Synonym: (tc)-t-pa. Engineered: yes. Two chain tissue plasminogen activator. Chain: b. Fragment: catalytic domain. Synonym: (tc)-t-pa.
Source: Homo sapiens. Human. Organism_taxid: 9606. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_taxid: 562
2.30Å     R-factor:   0.184     R-free:   0.265
Authors: W.Bode,D.Lamba
Key ref:
D.Lamba et al. (1996). The 2.3 A crystal structure of the catalytic domain of recombinant two-chain human tissue-type plasminogen activator. J Mol Biol, 258, 117-135. PubMed id: 8613982 DOI: 10.1006/jmbi.1996.0238
10-Nov-95     Release date:   11-Jan-97    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P00750  (TPA_HUMAN) -  Tissue-type plasminogen activator
562 a.a.
244 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: E.C.  - T-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     catalytic activity     2 terms  


DOI no: 10.1006/jmbi.1996.0238 J Mol Biol 258:117-135 (1996)
PubMed id: 8613982  
The 2.3 A crystal structure of the catalytic domain of recombinant two-chain human tissue-type plasminogen activator.
D.Lamba, M.Bauer, R.Huber, S.Fischer, R.Rudolph, U.Kohnert, W.Bode.
Tissue-type plasminogen activator (t-PA), a multidomainal serine proteinase of the trypsin-family, catalyses the rate-limiting step in fibrinolysis, the activation of plasminogen to the fibrin-degrading proteinase plasmin. Trigonal crystals have been obtained of the recombinant catalytic domain of human-two-chain t-PA, consisting of a 17 residue A chain and the 252 residue B chain. Its X-ray crystal structure has been solved applying Patterson and isomorphous replacement methods, and has been crystallographically refined to an R-value of 0.184 at 2.3 A resolution. The chain fold, active-site geometry and Ile276-Asp477 salt bridge are similar to that observed for trypsin. A few surface-located insertion loops differ significantly, however. The disulfide bridge Cys315-Cys384, practically unique to the plasminogen activators, is incorporated without drastic conformational changes as the insertion loop preceding Cys384 makes a bulge on the molecular surface. The unique basic insertion loop Lys296-Arg304 flanking the primed subsites, which has been shown to be of importance for PAI-1 binding and for fibrin specificity, is partially disordered; it can therefore freely adapt to proteins docking to the active site. The S1 pocket of t-PA is almost identical to that of trypsin, whereas the S2 site is considerably reduced in size by the imposing Tyr368 side-chain, in agreement with the measured preference for P1 Arg and P2 Gly residues. The neighbouring S3-S4 hydrophobic groove is mainly hydrophobic in nature. The structure of the proteinase domain of two-chain t-PA suggests that the formation of a salt bridge between Lys429 and Asp477 may contribute to the unusually high catalytic activity of single-chain t-PA, thus stabilizing the catalytically active conformation without unmasking the Ile276 amino terminus. Modeling studies show that the covalently bound kringle 2 domain in full-length t-PA could interact with an extended hydrophobic groove in the catalytic domain; in such a docking geometry its "lysine binding site" and the "fibrin binding patch" of the catalytic domain are in close proximity.
  Selected figure(s)  
Figure 3.
Figure 3. Stereo section of the final electron density map (blue) around the bound benzamidine molecule (center), superimposed on the t-PA model. Standard view as in Figures 1 and 2. Of the protein structure, only the entrance frame Trpc215 to Cysc220 (around the benzamidine), the catalytic triad Serc195, Hisc57 and Aspc102 (to the east), and Lysc143 and Tyrc151 (to the south) are displayed; the spherical density east of the benzamidine molecule, which partially hides the active Ser195, represents the bound phosphate ion. Contouring is at 1.0s. Figure made with O (Jones et al., 1991).
Figure 5.
Figure 5. Stereo plot of the hypothetical docking complex of the catalytic domain (white connections) and kringle 2 (yellow connections; De Vos et al., 1992) in the covalent two-domain t-PA variant. The domains are superimposed with a blue (catalytic domain) and a green Connolly surface (kringle 2 domain). This view is approximately rotated 135° from the standard orientation around a horizontal axis, so that the active site is now pointing to the east/back. Charged side-chains of catalytic domain residues presumably involved in fibrin binding, and kringle 2 residues forming the lysine binding site are labeled (chymotrypsinogen and kringle nomenclature). The plot was made with MAIN (Turk, 1992).
  The above figures are reprinted by permission from Elsevier: J Mol Biol (1996, 258, 117-135) copyright 1996.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21136135 J.Schaller, and S.S.Gerber (2011).
The plasmin-antiplasmin system: structural and functional aspects.
  Cell Mol Life Sci, 68, 785-801.  
19920152 M.A.Salameh, J.L.Robinson, D.Navaneetham, D.Sinha, B.J.Madden, P.N.Walsh, and E.S.Radisky (2010).
The amyloid precursor protein/protease nexin 2 Kunitz inhibitor domain is a highly specific substrate of mesotrypsin.
  J Biol Chem, 285, 1939-1949.  
17408411 C.Thelwell, and C.Longstaff (2007).
The regulation by fibrinogen and fibrin of tissue plasminogen activator kinetics and inhibition by plasminogen activator inhibitor 1.
  J Thromb Haemost, 5, 804-811.  
17996243 F.F.Liu, X.Y.Dong, T.Wang, and Y.Sun (2007).
Rational design of peptide ligand for affinity chromatography of tissue-type plasminogen activator by the combination of docking and molecular dynamics simulations.
  J Chromatogr A, 1175, 249-258.  
15516335 A.A.Komissarov, P.A.Andreasen, J.S.Bødker, P.J.Declerck, J.Y.Anagli, and J.D.Shore (2005).
Additivity in effects of vitronectin and monoclonal antibodies against alpha-helix F of plasminogen activator inhibitor-1 on its reactions with target proteinases.
  J Biol Chem, 280, 1482-1489.  
15892855 W.Bode (2005).
The structure of thrombin, a chameleon-like proteinase.
  J Thromb Haemost, 3, 2379-2388.  
16231330 Z.Sun, and J.N.Liu (2005).
Mutagenesis at Pro309 of single-chain urokinase-type plasminogen activator alters its catalytic properties.
  Proteins, 61, 870-877.  
15033993 A.A.Komissarov, P.J.Declerck, and J.D.Shore (2004).
Protonation state of a single histidine residue contributes significantly to the kinetics of the reaction of plasminogen activator inhibitor-1 with tissue-type plasminogen activator.
  J Biol Chem, 279, 23007-23013.  
14594804 C.A.Ibarra, G.E.Blouse, T.D.Christian, and J.D.Shore (2004).
The contribution of the exosite residues of plasminogen activator inhibitor-1 to proteinase inhibition.
  J Biol Chem, 279, 3643-3650.  
15331588 T.Tsuboi, H.T.McMahon, and G.A.Rutter (2004).
Mechanisms of dense core vesicle recapture following "kiss and run" ("cavicapture") exocytosis in insulin-secreting cells.
  J Biol Chem, 279, 47115-47124.  
14500731 M.J.Perron, G.E.Blouse, and J.D.Shore (2003).
Distortion of the catalytic domain of tissue-type plasminogen activator by plasminogen activator inhibitor-1 coincides with the formation of stable serpin-proteinase complexes.
  J Biol Chem, 278, 48197-48203.  
12468550 O.Roda, M.L.Valero, S.Peiró, D.Andreu, F.X.Real, and P.Navarro (2003).
New insights into the tPA-annexin A2 interaction. Is annexin A2 CYS8 the sole requirement for this association?
  J Biol Chem, 278, 5702-5709.  
12564979 Y.J.Park, J.F.Liang, H.Song, Y.T.Li, S.Naik, and V.C.Yang (2003).
ATTEMPTS: a heparin/protamine-based triggered release system for the delivery of enzyme drugs without associated side-effects.
  Adv Drug Deliv Rev, 55, 251-265.  
11772450 J.F.Liang, Y.T.Li, H.Song, Y.J.Park, S.S.Naik, and V.C.Yang (2002).
ATTEMPTS: a heparin/protamine-based delivery system for enzyme drugs.
  J Control Release, 78, 67-79.  
11967374 M.Backovic, E.Stratikos, D.A.Lawrence, and P.G.Gettins (2002).
Structural similarity of the covalent complexes formed between the serpin plasminogen activator inhibitor-1 and the arginine-specific proteinases trypsin, LMW u-PA, HMW u-PA, and t-PA: use of site-specific fluorescent probes of local environment.
  Protein Sci, 11, 1182-1191.  
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
11470437 C.Eigenbrot, D.Kirchhofer, M.S.Dennis, L.Santell, R.A.Lazarus, J.Stamos, and M.H.Ultsch (2001).
The factor VII zymogen structure reveals reregistration of beta strands during activation.
  Structure, 9, 627-636.
PDB code: 1jbu
11742690 K.Lähteenmäki, P.Kuusela, and T.K.Korhonen (2001).
Bacterial plasminogen activators and receptors.
  FEMS Microbiol Rev, 25, 531-552.  
  10752624 A.Vindigni, M.Winfield, Y.M.Ayala, and E.Di Cera (2000).
Role of residue Y99 in tissue plasminogen activator.
  Protein Sci, 9, 619-622.  
11080640 R.Tranter, J.A.Read, R.Jones, and R.L.Brady (2000).
Effector sites in the three-dimensional structure of mammalian sperm beta-acrosin.
  Structure, 8, 1179-1188.
PDB codes: 1fiw 1fiz
11086215 S.Braud, C.Bon, and A.Wisner (2000).
Snake venom proteins acting on hemostasis.
  Biochimie, 82, 851-859.  
10636881 S.Braud, M.A.Parry, R.Maroun, C.Bon, and A.Wisner (2000).
The contribution of residues 192 and 193 to the specificity of snake venom serine proteinases.
  J Biol Chem, 275, 1823-1828.  
10805774 S.Sperl, U.Jacob, N.Arroyo de Prada, J.Stürzebecher, O.G.Wilhelm, W.Bode, V.Magdolen, R.Huber, and L.Moroder (2000).
(4-aminomethyl)phenylguanidine derivatives as nonpeptidic highly selective inhibitors of human urokinase.
  Proc Natl Acad Sci U S A, 97, 5113-5118.
PDB code: 1ejn
10702294 V.Nienaber, J.Wang, D.Davidson, and J.Henkin (2000).
Re-engineering of human urokinase provides a system for structure-based drug design at high resolution and reveals a novel structural subsite.
  J Biol Chem, 275, 7239-7248.  
  10975580 Y.M.Ayala, and E.Di Cera (2000).
A simple method for the determination of individual rate constants for substrate hydrolysis by serine proteases.
  Protein Sci, 9, 1589-1593.  
11092938 Y.Tang, J.Zhang, L.Gui, C.Wu, R.Fan, W.Chang, and D.Liang (2000).
Crystallization and preliminary X-ray analysis of earthworm fibrinolytic enzyme component A from Eisenia fetida.
  Acta Crystallogr D Biol Crystallogr, 56, 1659-1661.  
10419460 F.Werner, T.M.Razzaq, and V.Ellis (1999).
Tissue plasminogen activator binds to human vascular smooth muscle cells by a novel mechanism. Evidence for a reciprocal linkage between inhibition of catalytic activity and cellular binding.
  J Biol Chem, 274, 21555-21561.  
10102985 H.Czapinska, and J.Otlewski (1999).
Structural and energetic determinants of the S1-site specificity in serine proteases.
  Eur J Biochem, 260, 571-595.  
10467148 K.P.Hopfner, A.Lang, A.Karcher, K.Sichler, E.Kopetzki, H.Brandstetter, R.Huber, W.Bode, and R.A.Engh (1999).
Coagulation factor IXa: the relaxed conformation of Tyr99 blocks substrate binding.
  Structure, 7, 989-996.
PDB code: 1rfn
10497153 M.M.Krem, T.Rose, and E.Di Cera (1999).
The C-terminal sequence encodes function in serine proteases.
  J Biol Chem, 274, 28063-28066.  
10066774 Y.L.Zhang, L.Hervio, L.Strandberg, and E.L.Madison (1999).
Distinct contributions of residue 192 to the specificity of coagulation and fibrinolytic serine proteases.
  J Biol Chem, 274, 7153-7156.  
9819212 A.Pasternak, X.Liu, T.Y.Lin, and L.Hedstrom (1998).
Activating a zymogen without proteolytic processing: mutation of Lys15 and Asn194 activates trypsinogen.
  Biochemistry, 37, 16201-16210.  
  9568890 A.R.Khan, and M.N.James (1998).
Molecular mechanisms for the conversion of zymogens to active proteolytic enzymes.
  Protein Sci, 7, 815-836.  
  10082369 A.Vindigni, and E.Di Cera (1998).
Role of P225 and the C136-C201 disulfide bond in tissue plasminogen activator.
  Protein Sci, 7, 1728-1737.  
9789329 C.Birchmeier, and E.Gherardi (1998).
Developmental roles of HGF/SF and its receptor, the c-Met tyrosine kinase.
  Trends Cell Biol, 8, 404-410.  
9751643 G.S.Coombs, R.C.Bergstrom, J.L.Pellequer, S.I.Baker, M.Navre, M.M.Smith, J.A.Tainer, E.L.Madison, and D.R.Corey (1998).
Substrate specificity of prostate-specific antigen (PSA).
  Chem Biol, 5, 475-488.
PDB code: 2psa
9799512 J.O.Kvassman, I.Verhamme, and J.D.Shore (1998).
Inhibitory mechanism of serpins: loop insertion forces acylation of plasminogen activator by plasminogen activator inhibitor-1.
  Biochemistry, 37, 15491-15502.  
9783753 M.A.Parry, C.Fernandez-Catalan, A.Bergner, R.Huber, K.P.Hopfner, B.Schlott, K.H.Gührs, and W.Bode (1998).
The ternary microplasmin-staphylokinase-microplasmin complex is a proteinase-cofactor-substrate complex in action.
  Nat Struct Biol, 5, 917-923.
PDB code: 1bui
9753698 M.A.Parry, U.Jacob, R.Huber, A.Wisner, C.Bon, and W.Bode (1998).
The crystal structure of the novel snake venom plasminogen activator TSV-PA: a prototype structure for snake venom serine proteinases.
  Structure, 6, 1195-1206.
PDB code: 1bqy
9733510 X.Wang, X.Lin, J.A.Loy, J.Tang, and X.C.Zhang (1998).
Crystal structure of the catalytic domain of human plasmin complexed with streptokinase.
  Science, 281, 1662-1665.
PDB code: 1bml
9485445 Z.Sun, B.F.Liu, Y.Chen, V.Gurewich, D.Zhu, and J.N.Liu (1998).
Analysis of the forces which stabilize the active conformation of urokinase-type plasminogen activator.
  Biochemistry, 37, 2935-2940.  
8995220 K.Tachias, and E.L.Madison (1997).
Converting tissue type plasminogen activator into a zymogen. Important role of Lys156.
  J Biol Chem, 272, 28-31.  
9354616 M.Renatus, M.T.Stubbs, R.Huber, P.Bringmann, P.Donner, W.D.Schleuning, and W.Bode (1997).
Catalytic domain structure of vampire bat plasminogen activator: a molecular paradigm for proteolysis without activation cleavage.
  Biochemistry, 36, 13483-13493.
PDB code: 1a5i
9305622 M.Renatus, R.A.Engh, M.T.Stubbs, R.Huber, S.Fischer, U.Kohnert, and W.Bode (1997).
Lysine 156 promotes the anomalous proenzyme activity of tPA: X-ray crystal structure of single-chain human tPA.
  EMBO J, 16, 4797-4805.
PDB code: 1bda
9268299 M.Renatus, W.Bode, R.Huber, J.Stürzebecher, D.Prasa, S.Fischer, U.Kohnert, and M.T.Stubbs (1997).
Structural mapping of the active site specificity determinants of human tissue-type plasminogen activator. Implications for the design of low molecular weight substrates and inhibitors.
  J Biol Chem, 272, 21713-21719.
PDB code: 1a5h
9047311 S.Butenas, M.Kalafatis, and K.G.Mann (1997).
Analysis of tissue plasminogen activator specificity using peptidyl fluorogenic substrates.
  Biochemistry, 36, 2123-2131.  
9252355 S.H.Ke, G.S.Coombs, K.Tachias, D.R.Corey, and E.L.Madison (1997).
Optimal subsite occupancy and design of a selective inhibitor of urokinase.
  J Biol Chem, 272, 20456-20462.  
8999865 S.H.Ke, K.Tachias, D.Lamba, W.Bode, and E.L.Madison (1997).
Identification of a hydrophobic exosite on tissue type plasminogen activator that modulates specificity for plasminogen.
  J Biol Chem, 272, 1811-1816.  
9434908 W.Bode, and M.Renatus (1997).
Tissue-type plasminogen activator: variants and crystal/solution structures demarcate structural determinants of function.
  Curr Opin Struct Biol, 7, 865-872.  
9252366 Y.Zhang, A.Wisner, R.C.Maroun, V.Choumet, Y.Xiong, and C.Bon (1997).
Trimeresurus stejnegeri snake venom plasminogen activator. Site-directed mutagenesis and molecular modeling.
  J Biol Chem, 272, 20531-20537.  
9295329 Z.Sun, Y.Jiang, Z.Ma, H.Wu, B.F.Liu, Y.Xue, W.Tang, Y.Chen, C.Li, D.Zhu, V.Gurewich, J.N.Liu, M.Zhong, and Y.Xu (1997).
Identification of a flexible loop region (297-313) of urokinase-type plasminogen activator, which helps determine its catalytic activity.
  J Biol Chem, 272, 23818-23823.  
8962059 C.D.Dickinson, C.R.Kelly, and W.Ruf (1996).
Identification of surface residues mediating tissue factor binding and catalytic function of the serine protease factor VIIa.
  Proc Natl Acad Sci U S A, 93, 14379-14384.  
8910515 K.Tachias, and E.L.Madison (1996).
Converting tissue-type plasminogen activator into a zymogen.
  J Biol Chem, 271, 28749-28752.  
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.