PDBsum entry 2w5v

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Hydrolase PDB id
Protein chains
346 a.a. *
_MG ×4
_ZN ×4
Waters ×589
* Residue conservation analysis
PDB id:
Name: Hydrolase
Title: Structure of tab5 alkaline phosphatase mutant his 135 asp with mg bound in the m3 site.
Structure: Alkaline phosphatase. Chain: a, b. Synonym: tab5 alkaline phosphatase mutant. Engineered: yes. Mutation: yes. Other_details: phosphoserine residue at position 84. M1, m2 occupied by zn, m3 by mg.
Source: Antarctic bacterium tab5. Organism_taxid: 82349. Expressed in: escherichia coli. Expression_system_taxid: 562
1.78Å     R-factor:   0.161     R-free:   0.199
Authors: D.Koutsioulis,A.Lyskowski,S.Maki,E.Guthrie,G.Feller, V.Bouriotis,P.Heikinheimo
Key ref: D.Koutsioulis et al. (2010). Coordination sphere of the third metal site is essential to the activity and metal selectivity of alkaline phosphatases. Protein Sci, 19, 75-84. PubMed id: 19916164
15-Dec-08     Release date:   24-Nov-09    
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Protein chains
Pfam   ArchSchema ?
Q9KWY4  (Q9KWY4_9BACT) -  Alkaline phosphatase
375 a.a.
346 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 3 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     metabolic process   2 terms 
  Biochemical function     catalytic activity     3 terms  


Protein Sci 19:75-84 (2010)
PubMed id: 19916164  
Coordination sphere of the third metal site is essential to the activity and metal selectivity of alkaline phosphatases.
D.Koutsioulis, A.Lyskowski, S.Mäki, E.Guthrie, G.Feller, V.Bouriotis, P.Heikinheimo.
Alkaline phosphatases (APs) are commercially applied enzymes that catalyze the hydrolysis of phosphate monoesters by a reaction involving three active site metal ions. We have previously identified H135 as the key residue for controlling activity of the psychrophilic TAB5 AP (TAP). In this article, we describe three X-ray crystallographic structures on TAP variants H135E and H135D in complex with a variety of metal ions. The structural analysis is supported by thermodynamic and kinetic data. The AP catalysis essentially requires octahedral coordination in the M3 site, but stability is adjusted with the conformational freedom of the metal ion. Comparison with the mesophilic Escherichia coli, AP shows differences in the charge transfer network in providing the chemically optimal metal combination for catalysis. Our results provide explanation why the TAB5 and E. coli APs respond in an opposite way to mutagenesis in their active sites. They provide a lesson on chemical fine tuning and the importance of the second coordination sphere in defining metal specificity in enzymes. Understanding the framework of AP catalysis is essential in the efforts to design even more powerful tools for modern biotechnology.