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PDBsum entry 3lb9

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protein Protein-protein interface(s) links
Hydrolase PDB id
3lb9
Jmol
Contents
Protein chains
182 a.a. *
Waters ×40
* Residue conservation analysis
PDB id:
3lb9
Name: Hydrolase
Title: Crystal structure of the b. Circulans cpa123 circular permut
Structure: Endo-1,4-beta-xylanase. Chain: a, b, c. Fragment: residues 65-182 and 2-63. Synonym: xylanase, 1,4-beta-d-xylan xylanohydrolase. Engineered: yes
Source: Bacillus circulans. Organism_taxid: 1397. Gene: xlna. Expressed in: escherichia coli. Expression_system_taxid: 469008.
Resolution:
3.00Å     R-factor:   0.195     R-free:   0.268
Authors: I.D'Angelo,S.Reitinger,M.Ludwiczek,N.Strynadka,S.G.Withers, L.P.Mcintosh
Key ref: S.Reitinger et al. (2010). Circular permutation of Bacillus circulans xylanase: a kinetic and structural study. Biochemistry, 49, 2464-2474. PubMed id: 20163191 DOI: 10.1021/bi100036f
Date:
08-Jan-10     Release date:   23-Mar-10    
PROCHECK
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 Headers
 References

Protein chains
Pfam   ArchSchema ?
P09850  (XYNA_BACCI) -  Endo-1,4-beta-xylanase
Seq:
Struc:
213 a.a.
182 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 25 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.3.2.1.8  - Endo-1,4-beta-xylanase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Endohydrolysis of 1,4-beta-D-xylosidic linkages in xylans.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     carbohydrate metabolic process   1 term 
  Biochemical function     hydrolase activity, hydrolyzing O-glycosyl compounds     1 term  

 

 
DOI no: 10.1021/bi100036f Biochemistry 49:2464-2474 (2010)
PubMed id: 20163191  
 
 
Circular permutation of Bacillus circulans xylanase: a kinetic and structural study.
S.Reitinger, Y.Yu, J.Wicki, M.Ludwiczek, I.D'Angelo, S.Baturin, M.Okon, N.C.Strynadka, S.Lutz, S.G.Withers, L.P.McIntosh.
 
  ABSTRACT  
 
The 20 kDa Bacillus circulans Bcx is a well-studied endoxylanase with a beta-jellyroll fold that places its N- and C-termini in salt bridge contact. Initial experiments verified that Bcx could be circularly permuted by PCR methods to introduce new termini in loop regions while linking its native termini directly or via one or two glycines. Subsequently, a library of circular permutants, generated by random DNase cleavage of the circularized Bcx gene, was screened for xylanase activity on xylan in Congo Red-stained agar. Analysis of 35 unique active circular permutants revealed that, while many of the new termini were introduced in external loops as anticipated, a surprising number were also located within beta-strands. Furthermore, several permutations placed key catalytic residues at or near the new termini with minimal deleterious effects on activity and, in one case, a 4-fold increase. The structure of one permutant was determined by X-ray crystallography, whereas three others were probed by NMR spectroscopy. These studies revealed that the overall conformation of Bcx changed very little in response to circular permutation, with effects largely being limited to increased local mobility near the new and the linked old termini and to a decrease in global stability against thermal denaturation. This library of circularly permuted xylanases provides an excellent set of new start points for directed evolution of this commercially important enzyme, as well as valuable constructs for intein-mediated replacement of key catalytic residues with unnatural analogues. Such approaches should permit new insights into the mechanism of enzymatic glycoside hydrolysis.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
21087800 Y.Yu, and S.Lutz (2011).
Circular permutation: a different way to engineer enzyme structure and function.
  Trends Biotechnol, 29, 18-25.  
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