A common structural basis for pH- and calmodulin-mediated regulation in plant glutamate decarboxylase.
Glutamate decarboxylase (Gad) catalyzes glutamate to gamma-aminobutyrate conversion. Plant Gad is a approximately 340 kDa hexamer, involved in development and stress response, and regulated by pH and binding of Ca(2+)/calmodulin (CaM) to the C-terminal domain. We determined the crystal structure of Arabidopsis thaliana Gad1 in its CaM-free state, obtained a low-resolution structure of the calmodulin-activated Gad complex by small-angle X-ray scattering and identified the crucial residues, in the C-terminal domain, for regulation by pH and CaM binding. CaM activates Gad1 in a unique way by relieving two C-terminal autoinhibition domains of adjacent active sites, forming a 393 kDa Gad1-CaM complex with an unusual 1:3 stoichiometry. The complex is loosely packed: thanks to the flexible linkers connecting the enzyme core with the six C-terminal regulatory domains, the CaM molecules retain considerable positional and orientational freedom with respect to Gad1. The complex thus represents a prototype for a novel CaM-target interaction mode. Thanks to its two levels of regulation, both targeting the C-terminal domain, Gad can respond flexibly to different kinds of cellular stress occurring at different pH values.
Gut
Heinz
H
Biochemisches Institut, Universität Zürich, Zürich, Switzerland.
Dominici
Paola
P
Pilati
Stefania
S
Astegno
Alessandra
A
Petoukhov
Maxim V
MV
Svergun
Dmitri I
DI
Grütter
Markus G
MG
Capitani
Guido
G
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000737:chemistry / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism*; D018360:Crystallography, X-Ray; D005968:Glutamate Decarboxylase / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism*; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D053838:Scattering, Small Angle; D016415:Sequence Alignment
D002147:Calmodulin; D010940:Plant Proteins; D005968:Glutamate Decarboxylase