Reference : Coordination sphere of the third metal site is essential to the activity and metal se...
Scientific journals : Article
Life sciences : Biochemistry, biophysics & molecular biology
Coordination sphere of the third metal site is essential to the activity and metal selectivity of alkaline phosphatases
Koutsioulis, D. [> > > >]
Lyskowski, A. [> > > >]
Maki, S. [> > > >]
Guthrie, E. [> > > >]
Feller, Georges mailto [Université de Liège - ULg > Département des sciences de la vie > Labo de biochimie >]
Bouriotis, V. [> > > >]
Heikinheimo, P. [> > > >]
Protein Science : A Publication of the Protein Society
Cold Spring Harbor Laboratory Press
Yes (verified by ORBi)
[en] 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.

File(s) associated to this reference

Fulltext file(s):

Open access
ProtSci_2010_AP.pdfAuthor postprint283.87 kBView/Open

Bookmark and Share SFX Query

All documents in ORBi are protected by a user license.