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PDBsum entry 1ejm
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protein ligands Protein-protein interface(s) links
Hydrolase/inhibitor PDB id
1ejm

 

 

 

 

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Contents
Protein chains
223 a.a. *
58 a.a. *
Ligands
SO4 ×12
Waters ×611
* Residue conservation analysis
PDB id:
1ejm
Name: Hydrolase/inhibitor
Title: Crystal structure of the bpti ala16leu mutant in complex with bovine trypsin
Structure: Beta-trypsin. Chain: a, c, e. Pancreatic trypsin inhibitor. Chain: b, d, f. Synonym: basic protease inhibitor, bpi, aprotinin. Engineered: yes. Mutation: yes
Source: Bos taurus. Cattle. Organism_taxid: 9913. Organ: pancreas. Other_details: purchased from sigma. Expressed in: escherichia coli. Expression_system_taxid: 562
Biol. unit: Tetramer (from PQS)
Resolution:
1.85Å     R-factor:   0.211     R-free:   0.233
Authors: J.Otlewski,A.Smalas,R.Helland,A.Grzesiak,D.Krowarsch
Key ref:
A.Grzesiak et al. (2000). Substitutions at the P(1) position in BPTI strongly affect the association energy with serine proteinases. J Mol Biol, 301, 205-217. PubMed id: 10926503 DOI: 10.1006/jmbi.2000.3935
Date:
03-Mar-00     Release date:   03-Mar-01    
PROCHECK
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 Headers
 References

Protein chains
Pfam   ArchSchema ?
P00760  (TRY1_BOVIN) -  Serine protease 1 from Bos taurus
Seq:
Struc: 		246 a.a.
223 a.a.
Protein chains
Pfam   ArchSchema ?
P00974  (BPT1_BOVIN) -  Pancreatic trypsin inhibitor from Bos taurus
Seq:
Struc: 		100 a.a.
58 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 3 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: Chains A, C, E: E.C.3.4.21.4  - trypsin.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Preferential cleavage: Arg-|-Xaa, Lys-|-Xaa.

 

 
DOI no: 10.1006/jmbi.2000.3935 J Mol Biol 301:205-217 (2000)
PubMed id: 10926503  
 
 
Substitutions at the P(1) position in BPTI strongly affect the association energy with serine proteinases.
A.Grzesiak, R.Helland, A.O.Smalås, D.Krowarsch, M.Dadlez, J.Otlewski.
 
  ABSTRACT  
 
The role of the S(1) subsite in trypsin, chymotrypsin and plasmin has been examined by measuring the association with seven different mutants of bovine pancreatic trypsin inhibitor (BPTI); the mutants contain Gly, Ala, Ser, Val, Leu, Arg, and Trp at the P(1) position of the reactive site. The effects of substitutions at the P(1) position on the association constants are very large, comprising seven orders of magnitude for trypsin and plasmin, and over five orders for chymotrypsin. All mutants showed a decrease of the association constant to the three proteinases in the same order: Ala>Gly>Ser>Arg>Val>Leu>Trp. Calorimetric and circular dichroism methods showed that none of the P1 substitutions, except the P1-Val mutant, lead to destabilisation of the binding loop conformation. The X-ray structure of the complex formed between bovine beta-trypsin and P(1)-Leu BPTI showed that the P(1)-Leu sterically conflicts with the side-chain of P(3)-Ile, which thereby is forced to rotate approximately 90 degrees. Ile18 (P(3)) in its new orientation, in turn interacts with the Tyr39 side-chain of trypsin. Introduction of a large side-chain at the P1' position apparently leads to a cascade of small alterations of the trypsin-BPTI interface that seem to destabilise the complex by it adopting a less optimized packing and by tilting the BPTI molecule up to 15 degrees compared to the native trypsin-BPTI complex.
 
  Selected figure(s)  
 
Figure 1.
Figure 1. Inhibition curves of human a-plasmin by Ala16, Ala16 -> Gly and Ala16 -> Arg mutants of BPTI. Reactions were carried out in 100 mM Tris-Cl, 20 mM CaCl[2], 150 mM NaCl, 0.05 % Triton X-100 (pH 8.3) at 298 K. Data points were analyzed using the equations described in Materials and Methods. 		Figure 7.
Figure 7. Stereo plot illustrating the competition between P'[1] and P'[3] residues for the same binding site. Red (inhibitor) and grey (trypsin) describes the bovine trypsin-BPTI P[1]-Lys complex while blue (inhibitor) and black (trypsin) describes the trypsin-BPTI P'[1]-Leu (molecule B) complex. The P[1], P'[1], P'[3] and P[4]' residues of the inhibitor are illustrated as ball and stick models. The Figures are generated using BOBSCRIPT [Kraulis 1991 and Esnouf 1997].
 
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2000, 301, 205-217) copyright 2000.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
22346912 G.Portalone (2012).
4-Meth-oxy-benzamidinium 2,6-dimeth-oxy-benzoate.
  Acta Crystallogr Sect E Struct Rep Online, 68, o268-o269.  
23284415 S.Irrera, and G.Portalone (2012).
4-Meth-oxy-benzamidinium chloride monohydrate.
  Acta Crystallogr Sect E Struct Rep Online, 68, o3083.  
23284542 S.Irrera, and G.Portalone (2012).
4-Meth-oxy-benzamidinium hydrogen sulfate.
  Acta Crystallogr Sect E Struct Rep Online, 68, o3244.  
20522949 G.Portalone (2010).
Supramolecular association in proton-transfer adducts containing benzamidinium cations. I. Four molecular salts with uracil derivatives.
  Acta Crystallogr C, 66, o295-o301.  
20597089 S.J.Fleishman, J.E.Corn, E.M.Strauch, T.A.Whitehead, I.Andre, J.Thompson, J.J.Havranek, R.Das, P.Bradley, and D.Baker (2010).
Rosetta in CAPRI rounds 13-19.
  Proteins, 78, 3212-3218.  
19490116 E.K.Millers, M.Trabi, P.P.Masci, M.F.Lavin, J.de Jersey, and L.W.Guddat (2009).
Crystal structure of textilinin-1, a Kunitz-type serine protease inhibitor from the venom of the Australian common brown snake (Pseudonaja textilis).
  FEBS J, 276, 3163-3175.
PDB code: 3byb
19435349 X.Li, X.He, B.Wang, and K.Merz (2009).
Conformational variability of benzamidinium-based inhibitors.
  J Am Chem Soc, 131, 7742-7754.  
18286181 S.Macedo-Ribeiro, C.Almeida, B.M.Calisto, T.Friedrich, R.Mentele, J.Stürzebecher, P.Fuentes-Prior, and P.J.Pereira (2008).
Isolation, cloning and structural characterisation of boophilin, a multifunctional Kunitz-type proteinase inhibitor from the cattle tick.
  PLoS ONE, 3, e1624.
PDB code: 2ody
16636277 E.S.Radisky, J.M.Lee, C.J.Lu, and D.E.Koshland (2006).
Insights into the serine protease mechanism from atomic resolution structures of trypsin reaction intermediates.
  Proc Natl Acad Sci U S A, 103, 6835-6840.
PDB codes: 2age 2agg 2agi 2ah4
16703468 F.J.Yan, C.P.Chen, Y.C.Cheng, and L.S.Chang (2006).
Mutagenesis studies on the N-terminus and Thr54 of Naja naja atra (Taiwan cobra) chymotrypsin inhibitor.
  Protein J, 25, 257-262.  
16755353 N.M.Talyzina, and P.K.Ingvarsson (2006).
Molecular evolution of a small gene family of wound inducible Kunitz trypsin inhibitors in Populus.
  J Mol Evol, 63, 108-119.  
15932872 A.E.Schmidt, H.S.Chand, D.Cascio, W.Kisiel, and S.P.Bajaj (2005).
Crystal structure of Kunitz domain 1 (KD1) of tissue factor pathway inhibitor-2 in complex with trypsin. Implications for KD1 specificity of inhibition.
  J Biol Chem, 280, 27832-27838.
PDB code: 1zr0
15578663 O.Guvench, D.J.Price, and C.L.Brooks (2005).
Receptor rigidity and ligand mobility in trypsin-ligand complexes.
  Proteins, 58, 407-417.  
15039345 D.Chu, R.D.Bungiro, M.Ibanez, L.M.Harrison, E.Campodonico, B.F.Jones, J.Mieszczanek, P.Kuzmic, and M.Cappello (2004).
Molecular characterization of Ancylostoma ceylanicum Kunitz-type serine protease inhibitor: evidence for a role in hookworm-associated growth delay.
  Infect Immun, 72, 2214-2221.  
12581670 M.Laskowski, M.A.Qasim, and Z.Yi (2003).
Additivity-based prediction of equilibrium constants for some protein-protein associations.
  Curr Opin Struct Biol, 13, 130-139.  
12384499 C.Hink-Schauer, E.Estébanez-Perpiñá, E.Wilharm, P.Fuentes-Prior, W.Klinkert, W.Bode, and D.E.Jenne (2002).
The 2.2-A crystal structure of human pro-granzyme K reveals a rigid zymogen with unusual features.
  J Biol Chem, 277, 50923-50933.
PDB codes: 1mza 1mzd
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 code is shown on the right.

 

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