PMID:12686133
| Citation |
James, PL and Anthony, C (2003) The metal ion in the active site of the membrane glucose dehydrogenase of Escherichia coli. Biochim. Biophys. Acta 1647:200-5 |
|---|---|
| Abstract |
All pyrroloquinoline quinone (PQQ)-containing dehydrogenases whose structures are known contain Ca(2+) bonded to the PQQ at the active site. However, membrane glucose dehydrogenase (GDH) requires reconstitution with PQQ and Mg(2+) ions (but not Ca(2+)) for activity. To address the question of whether the Mg(2+) replaces the usual active site Ca(2+) in this enzyme, mutant GDHs were produced in which residues proposed to be involved in binding metal ion were modified (D354N-GDH and N355D-GDH and D354N-GDH/N355D-GDH). The most remarkable observation was that reconstitution with PQQ of the mutant enzymes was not supported by Mg(2+) ions as in the wild-type GDH, but it could be supported by Ca(2+), Sr(2+) or Ba(2+) ions. This was competitively inhibited by Mg(2+). This result, together with studies on the kinetics of the modified enzymes have led to the conclusion that, although a Ca(2+) ion is able to form part of the active site of the genetically modified GDH, as in all other PQQ-containing quinoproteins, a Mg(2+) ion surprisingly replaces Ca(2+) in the active site of the wild-type GDH. |
| Links | |
| Keywords |
Binding Sites; Calcium/chemistry; Cell Membrane/enzymology; Escherichia coli/enzymology; Escherichia coli Proteins/chemistry; Glucose Dehydrogenases/chemistry; Magnesium/chemistry; Mutagenesis, Site-Directed; Substrate Specificity |
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