Crystal structure and functional analysis of Escherichia coli glutamate decarboxylase.
Glutamate decarboxylase is a vitamin B6-dependent enzyme, which catalyses the decarboxylation of glutamate to gamma-aminobutyrate. In Escherichia coli, expression of glutamate decarboxylase (GadB), a 330 kDa hexamer, is induced to maintain the physiological pH under acidic conditions, like those of the passage through the stomach en route to the intestine. GadB, together with the antiporter GadC, constitutes the gad acid resistance system, which confers the ability for bacterial survival for at least 2 h in a strongly acidic environment. GadB undergoes a pH-dependent conformational change and exhibits an activity optimum at low pH. We determined the crystal structures of GadB at acidic and neutral pH. They reveal the molecular details of the conformational change and the structural basis for the acidic pH optimum. We demonstrate that the enzyme is localized exclusively in the cytoplasm at neutral pH, but is recruited to the membrane when the pH falls. We show by structure-based site-directed mutagenesis that the triple helix bundle formed by the N-termini of the protein at acidic pH is the major determinant for this behaviour.
Capitani
Guido
G
Biochemisches Institut der Universität Zürich, Zürich CH-8057, Switzerland. capitani@bioc.unizh.ch
De Biase
Daniela
D
Aurizi
Caterina
C
Gut
Heinz
H
Bossa
Francesco
F
Grütter
Markus G
MG
D001665:Binding Sites; D018360:Crystallography, X-Ray; D019281:Dimerization; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D005968:Glutamate Decarboxylase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020449:Repetitive Sequences, Amino Acid; D019346:Sodium Acetate / Q000378:metabolism
D019346:Sodium Acetate; D005968:Glutamate Decarboxylase