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PDBsum entry 1kgw

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protein ligands metals links
Hydrolase PDB id
1kgw
Jmol
Contents
Protein chain
496 a.a. *
Ligands
NAG
Metals
_CA
Waters ×171
* Residue conservation analysis
PDB id:
1kgw
Name: Hydrolase
Title: Three dimensional structure analysis of the r337q variant of pancreatic alpha-mylase
Structure: Alpha-amylase, pancreatic. Chain: a. Synonym: 1,4-alpha-d-glucan glucanohydrolase, pancreatic al amylase, pa. Engineered: yes. Mutation: yes
Source: Homo sapiens. Human. Organism_taxid: 9606. Expressed in: pichia pastoris. Expression_system_taxid: 4922
Resolution:
2.10Å     R-factor:   0.190    
Authors: S.Numao,R.Maurus,G.Sidhu,Y.Wang,C.M.Overall,G.D.Brayer,S.G.W
Key ref:
S.Numao et al. (2002). Probing the role of the chloride ion in the mechanism of human pancreatic alpha-amylase. Biochemistry, 41, 215-225. PubMed id: 11772019 DOI: 10.1021/bi0115636
Date:
28-Nov-01     Release date:   16-Jan-02    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P04746  (AMYP_HUMAN) -  Pancreatic alpha-amylase
Seq:
Struc:
511 a.a.
496 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.3.2.1.1  - Alpha-amylase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Endohydrolysis of 1,4-alpha-glucosidic linkages in oligosaccharides and polysaccharides.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     extracellular region   3 terms 
  Biological process     metabolic process   5 terms 
  Biochemical function     catalytic activity     8 terms  

 

 
DOI no: 10.1021/bi0115636 Biochemistry 41:215-225 (2002)
PubMed id: 11772019  
 
 
Probing the role of the chloride ion in the mechanism of human pancreatic alpha-amylase.
S.Numao, R.Maurus, G.Sidhu, Y.Wang, C.M.Overall, G.D.Brayer, S.G.Withers.
 
  ABSTRACT  
 
Human pancreatic alpha-amylase (HPA) is a member of the alpha-amylase family involved in the degradation of starch. Some members of this family, including HPA, require chloride for maximal activity. To determine the mechanism of chloride activation, a series of mutants (R195A, R195Q, N298S, R337A, and R337Q) were made in which residues in the chloride ion binding site were replaced. Mutations in this binding site were found to severely affect the ability of HPA to bind chloride ions with no binding detected for the R195 and R337 mutant enzymes. X-ray crystallographic analysis revealed that these mutations did not result in significant structural changes. However, the introduction of these mutations did alter the kinetic properties of the enzyme. Mutations to residue R195 resulted in a 20-450-fold decrease in the activity of the enzyme toward starch and shifted the pH optimum to a more basic pH. Interestingly, replacement of R337 with a nonbasic amino acid resulted in an alpha-amylase that no longer required chloride for catalysis and has a pH profile similar to that of wild-type HPA. In contrast, a mutation at residue N298 resulted in an enzyme that had much lower binding affinity for chloride but still required chloride for maximal activity. We propose that the chloride is required to increase the pK(a) of the acid/base catalyst, E233, which would otherwise be lower due to the presence of R337, a positively charged residue.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
20461849 F.Cardona, A.Goti, C.Parmeggiani, P.Parenti, M.Forcella, P.Fusi, L.Cipolla, S.M.Roberts, G.J.Davies, and T.M.Gloster (2010).
Casuarine-6-O-alpha-D-glucoside and its analogues are tight binding inhibitors of insect and bacterial trehalases.
  Chem Commun (Camb), 46, 2629-2631.
PDB code: 2wyn
19476481 J.Pytelková, J.Hubert, M.Lepsík, J.Sobotník, R.Sindelka, I.Krízková, M.Horn, and M.Mares (2009).
Digestive alpha-amylases of the flour moth Ephestia kuehniella--adaptation to alkaline environment and plant inhibitors.
  FEBS J, 276, 3531-3546.  
18214874 C.A.Tarling, K.Woods, R.Zhang, H.C.Brastianos, G.D.Brayer, R.J.Andersen, and S.G.Withers (2008).
The search for novel human pancreatic alpha-amylase inhibitors: high-throughput screening of terrestrial and marine natural product extracts.
  Chembiochem, 9, 433-438.  
17729287 J.C.Marx, J.Poncin, J.P.Simorre, P.W.Ramteke, and G.Feller (2008).
The noncatalytic triad of alpha-amylases: a novel structural motif involved in conformational stability.
  Proteins, 70, 320-328.  
18613721 S.Cheluvaraja, M.Mihailescu, and H.Meirovitch (2008).
Entropy and free energy of a mobile protein loop in explicit water.
  J Phys Chem B, 112, 9512-9522.  
17359918 A.J.Plested, and M.L.Mayer (2007).
Structure and mechanism of kainate receptor modulation by anions.
  Neuron, 53, 829-841.
PDB code: 2ojt
15722449 R.Maurus, A.Begum, H.H.Kuo, A.Racaza, S.Numao, C.Andersen, J.W.Tams, J.Vind, C.M.Overall, S.G.Withers, and G.D.Brayer (2005).
Structural and mechanistic studies of chloride induced activation of human pancreatic alpha-amylase.
  Protein Sci, 14, 743-755.
PDB codes: 1xgz 1xh0 1xh1 1xh2
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.