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PDBsum entry 1pj9

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protein ligands metals links
Transferase PDB id
1pj9
Jmol
Contents
Protein chain
686 a.a. *
Ligands
MAL
GLC-GLC ×2
GLC-GLC-GLC ×2
GLC
MPD
ACY ×3
Metals
_CA ×3
Waters ×798
* Residue conservation analysis
PDB id:
1pj9
Name: Transferase
Title: Bacillus circulans strain 251 loop mutant 183-195
Structure: Cyclomaltodextrin glucanotransferase. Chain: a. Synonym: cyclodextrin-glycosyltransferase, cgtase. Engineered: yes. Mutation: yes
Source: Bacillus circulans. Organism_taxid: 1397. Strain: 251. Expressed in: escherichia coli. Expression_system_taxid: 562.
Resolution:
2.00Å     R-factor:   0.152     R-free:   0.176
Authors: H.J.Rozeboom,B.W.Dijkstra
Key ref:
H.Leemhuis et al. (2004). Improved thermostability of bacillus circulans cyclodextrin glycosyltransferase by the introduction of a salt bridge. Proteins, 54, 128-134. PubMed id: 14705029 DOI: 10.1002/prot.10516
Date:
02-Jun-03     Release date:   03-Feb-04    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P43379  (CDGT2_BACCI) -  Cyclomaltodextrin glucanotransferase
Seq:
Struc:
 
Seq:
Struc:
713 a.a.
686 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 6 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.2.4.1.19  - Cyclomaltodextrin glucanotransferase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Degrades starch to cyclodextrins by formation of a 1,4-alpha-D- glucosidic bond.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     extracellular region   1 term 
  Biological process     carbohydrate metabolic process   1 term 
  Biochemical function     catalytic activity     8 terms  

 

 
DOI no: 10.1002/prot.10516 Proteins 54:128-134 (2004)
PubMed id: 14705029  
 
 
Improved thermostability of bacillus circulans cyclodextrin glycosyltransferase by the introduction of a salt bridge.
H.Leemhuis, H.J.Rozeboom, B.W.Dijkstra, L.Dijkhuizen.
 
  ABSTRACT  
 
Cyclodextrin glycosyltransferase (CGTase) catalyzes the formation of cyclodextrins from starch. Among the CGTases with known three-dimensional structure, Thermoanaerobacterium thermosulfurigenes CGTase has the highest thermostability. By replacing amino acid residues in the B-domain of Bacillus circulans CGTase with those from T. thermosulfurigenes CGTase, we identified a B. circulans CGTase mutant (with N188D and K192R mutations), with a strongly increased activity half-life at 60 degrees C. Asp188 and Arg192 form a salt bridge in T. thermosulfurigenes CGTase. Structural analysis of the B. circulans CGTase mutant revealed that this salt bridge is also formed in the mutant. Thus, the activity half-life of this enzyme can be enhanced by rational protein engineering.
 
  Selected figure(s)  
 
Figure 1.
Figure 1. Close-up views of the loop region 185-192 in CGTase. For clarity, only the backbone and the sidechains of residues 188 and 192 are shown (Asn/Lys in BC251 CGTase; Asp/Arg in Tabium, B. stearothermophilus, and mutant 9 CGTases). (A) BC251 and Tabium CGTase, (B) BC251 and mutant 9 CGTase, (C) Tabium and mutant 9 CGTase, and (D) mutant 9 and B. stearothermophilus CGTase. Salt bridges are indicated by dashed lines. The CGTases are shown in black (BC251 and B. stearothermophilus), gray (Tabium), and light gray (mutant 9).
Figure 2.
Figure 2. DSC curves for the thermal denaturation of wild-type BC251 CGTase (1), BC251 CGTase mutant 9 (2), and Tabium CGTase (3), in the absence (A) and presence (B) of 10 mM added Ca^2+ ions.
 
  The above figures are reprinted by permission from John Wiley & Sons, Inc.: Proteins (2004, 54, 128-134) copyright 2004.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19763564 H.Leemhuis, R.M.Kelly, and L.Dijkhuizen (2010).
Engineering of cyclodextrin glucanotransferases and the impact for biotechnological applications.
  Appl Microbiol Biotechnol, 85, 823-835.  
19367403 R.M.Kelly, L.Dijkhuizen, and H.Leemhuis (2009).
The evolution of cyclodextrin glucanotransferase product specificity.
  Appl Microbiol Biotechnol, 84, 119-133.  
20054483 S.Wang, Y.B.Yan, and Z.Y.Dong (2009).
Contributions of the C-Terminal Helix to the Structural Stability of a Hyperthermophilic Fe-Superoxide Dismutase (TcSOD).
  Int J Mol Sci, 10, 5498-5512.  
18283101 R.M.Kelly, H.Leemhuis, L.Gätjen, and L.Dijkhuizen (2008).
Evolution toward small molecule inhibitor resistance affects native enzyme function and stability, generating acarbose-insensitive cyclodextrin glucanotransferase variants.
  J Biol Chem, 283, 10727-10734.  
18726621 R.M.Ong, K.M.Goh, N.M.Mahadi, O.Hassan, R.N.Rahman, and R.M.Illias (2008).
Cloning, extracellular expression and characterization of a predominant beta-CGTase from Bacillus sp. G1 in E. coli.
  J Ind Microbiol Biotechnol, 35, 1705-1714.  
17968540 W.Zhang, and X.G.Lei (2008).
Cumulative improvements of thermostability and pH-activity profile of Aspergillus niger PhyA phytase by site-directed mutagenesis.
  Appl Microbiol Biotechnol, 77, 1033-1040.  
16247799 Ihsanawati, T.Kumasaka, T.Kaneko, C.Morokuma, R.Yatsunami, T.Sato, S.Nakamura, and N.Tanaka (2005).
Structural basis of the substrate subsite and the highly thermal stability of xylanase 10B from Thermotoga maritima MSB8.
  Proteins, 61, 999.
PDB codes: 1vbr 1vbu
15522513 M.Díaz, S.Rodriguez, J.M.Fernández-Abalos, J.De Las Rivas, A.Ruiz-Arribas, V.L.Shnyrov, and R.I.Santamaría (2004).
Single mutations of residues outside the active center of the xylanase Xys1 Delta from Streptomyces halstedii JM8 affect its activity.
  FEMS Microbiol Lett, 240, 237-243.  
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.