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PDBsum entry 2abz

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protein metals Protein-protein interface(s) links
Hydrolase/hydrolase inhibitor PDB id
2abz
Jmol
Contents
Protein chains
299 a.a. *
62 a.a. *
46 a.a. *
60 a.a. *
58 a.a. *
Metals
_ZN ×2
* Residue conservation analysis
PDB id:
2abz
Name: Hydrolase/hydrolase inhibitor
Title: Crystal structure of c19a/c43a mutant of leech carboxypeptidase inhibitor in complex with bovine carboxypeptidase a
Structure: Carboxypeptidase a1. Chain: a, b. Synonym: carboxypeptidase a, cpa, a/b metallocarboxypeptidase. Engineered: yes. Metallocarboxypeptidase inhibitor. Chain: c, d, e, f. Synonym: leech carboxypeptidase inhibitor, lci, inhibitor of a/b metallocarboxypeptidases.
Source: Bos taurus. Cattle. Organism_taxid: 9913. Expressed in: pichia pastoris. Expression_system_taxid: 4922. Hirudo medicinalis. Medicinal leech. Organism_taxid: 6421. Expressed in: escherichia coli bl21(de3).
Biol. unit: Hexamer (from PQS)
Resolution:
2.16Å     R-factor:   0.190     R-free:   0.234
Authors: J.L.Arolas,G.M.Popowicz,S.Bronsoms,F.X.Aviles,R.Huber, T.A.Holak,S.Ventura
Key ref:
J.L.Arolas et al. (2005). Study of a major intermediate in the oxidative folding of leech carboxypeptidase inhibitor: contribution of the fourth disulfide bond. J Mol Biol, 352, 961-975. PubMed id: 16126224 DOI: 10.1016/j.jmb.2005.07.065
Date:
18-Jul-05     Release date:   31-Jan-06    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P00730  (CBPA1_BOVIN) -  Carboxypeptidase A1
Seq:
Struc:
419 a.a.
299 a.a.*
Protein chain
Pfam   ArchSchema ?
P81511  (MCPI_HIRME) -  Metallocarboxypeptidase inhibitor
Seq:
Struc:
81 a.a.
62 a.a.*
Protein chain
No UniProt id for this chain
Struc: 46 a.a.
Protein chain
Pfam   ArchSchema ?
P81511  (MCPI_HIRME) -  Metallocarboxypeptidase inhibitor
Seq:
Struc:
81 a.a.
60 a.a.*
Protein chain
Pfam   ArchSchema ?
P81511  (MCPI_HIRME) -  Metallocarboxypeptidase inhibitor
Seq:
Struc:
81 a.a.
58 a.a.*
Key:    PfamA domain  PfamB domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 8 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: Chains A, B: E.C.3.4.17.1  - Carboxypeptidase A.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Peptidyl-L-amino acid + H2O = peptide + L-amino acid

+
=
+
      Cofactor: Zinc
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     proteolysis   3 terms 
  Biochemical function     enzyme inhibitor activity     5 terms  

 

 
    Added reference    
 
 
DOI no: 10.1016/j.jmb.2005.07.065 J Mol Biol 352:961-975 (2005)
PubMed id: 16126224  
 
 
Study of a major intermediate in the oxidative folding of leech carboxypeptidase inhibitor: contribution of the fourth disulfide bond.
J.L.Arolas, G.M.Popowicz, S.Bronsoms, F.X.Aviles, R.Huber, T.A.Holak, S.Ventura.
 
  ABSTRACT  
 
The oxidative folding pathway of leech carboxypeptidase inhibitor (LCI; four disulfide bonds) proceeds through the formation of two major intermediates (III-A and III-B) that contain three native disulfide bonds and act as strong kinetic traps in the folding process. The III-B intermediate lacks the Cys19-Cys43 disulfide bond that links the beta-sheet core with the alpha-helix in wild-type LCI. Here, an analog of this intermediate was constructed by replacing Cys19 and Cys43 with alanine residues. Its oxidative folding follows a rapid sequential flow through one, two, and three disulfide species to reach the native form; the low accumulation of two disulfide intermediates and three disulfide (scrambled) isomers accounts for a highly efficient reaction. The three-dimensional structure of this analog, alone and in complex with carboxypeptidase A (CPA), was determined by X-ray crystallography at 2.2A resolution. Its overall structure is very similar to that of wild-type LCI, although the residues in the region adjacent to the mutation sites show an increased flexibility, which is strongly reduced upon binding to CPA. The structure of the complex also demonstrates that the analog and the wild-type LCI bind to the enzyme in the same manner, as expected by their inhibitory capabilities, which were similar for all enzymes tested. Equilibrium unfolding experiments showed that this mutant is destabilized by approximately 1.5 kcal mol(-1) (40%) relative to the wild-type protein. Together, the data indicate that the fourth disulfide bond provides LCI with both high stability and structural specificity.
 
  Selected figure(s)  
 
Figure 7.
Figure 7. Three-dimensional structure of C19A/C43A LCI. (a) Stereo representation of the C19A/C43A LCI ribbon. The helix (a1) and b-strands (b1-b5) are represented in red and light blue, respectively. The three disulfide bridges of this mutant (Cys11-Cys34, Cys22-Cys58 and Cys18-Cys62) are represented in yellow. N and C indicate the location of N and C-terminal tails of C19A/C43A LCI. (b) Stereo view of the overlapping between the backbone atoms from the bound form of C19A/C43A (green) and wt LCI (orange). The disulfide pairings of each protein are shown yellow in the structure. (c) Stereo view of the overlapping between the backbone atoms from the free (red) and bound (green) forms of C19A/C43A LCI. The disulfide pairings are indicated in yellow.
Figure 8.
Figure 8. Stereo ribbon representation of the complex formed between C19A/C43A LCI and CPA. The helix and b-strands of C19A/C43A LCI are shown in red and light blue, respectively, and the disulfide bridges are in yellow. The side-chains of C19A/C43A LCI residues involved in the interaction with CPA are explicitly shown in orange. CPA is colored green and its catalytic zinc atom is represented by a yellow sphere. The N-terminal and C-terminal domains of the mutant and the enzyme are depicted.
 
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2005, 352, 961-975) copyright 2005.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20812859 J.L.Arolas, and S.Ventura (2011).
Protease inhibitors as models for the study of oxidative folding.
  Antioxid Redox Signal, 14, 97.  
17961067 J.Y.Chang (2008).
Diversity of folding pathways and folding models of disulfide proteins.
  Antioxid Redox Signal, 10, 171-178.  
18547517 M.Cemazar, A.Joshi, N.L.Daly, A.E.Mark, and D.J.Craik (2008).
The structure of a two-disulfide intermediate assists in elucidating the oxidative folding pathway of a cyclic cystine knot protein.
  Structure, 16, 842-851.  
16600598 J.L.Arolas, F.X.Aviles, J.Y.Chang, and S.Ventura (2006).
Folding of small disulfide-rich proteins: clarifying the puzzle.
  Trends Biochem Sci, 31, 292-301.  
16710754 S.Salamanca, and J.Y.Chang (2006).
Pathway of oxidative folding of a 3-disulfide alpha-lactalbumin may resemble either BPTI model or hirudin model.
  Protein J, 25, 275-287.  
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.