A Common Structural Basis for pH- and Calmodulin-mediated Regulation in Plant Glutamate Decarboxylase

Gut, H.; Dominici, Paola; Pilati, S.; Astegno, Alessandra; Petoukhov, Maxim V.; Svergun, Dmitri I.; Grütter, Markus G.; Capitani, Guido

doi:10.1016/j.jmb.2009.06.080

Cited by 76 publications

(80 citation statements)

References 66 publications

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“…While our structural studies suggest the AQP0–CaM interaction is similar to the pt GAD–CaM interaction, the regulatory mechanisms are quite different. For pt GAD, CaM binding relieves an auto-inhibitory domain of the enzyme that results in functional activation 54 . For AQP0, CaM binding induces allosteric changes in the channel gate that restrict water permeation.…”

Section: Discussionmentioning

confidence: 99%

Allosteric mechanism of water-channel gating by Ca2+–calmodulin

Reichow

Clemens

Freites

et al. 2013

Nat Struct Mol Biol

106

138

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Calmodulin (CaM) is a universal regulatory protein that communicates the presence of calcium to its molecular targets and correspondingly modulates their function. This key signaling protein is important for controlling the activity of hundreds of membrane channels and transporters. However, our understanding of the structural mechanisms driving CaM regulation of full-length membrane proteins has remained elusive. In this study, we determined the pseudo-atomic structure of full-length mammalian aquaporin-0 (AQP0, Bos Taurus) in complex with CaM using electron microscopy to understand how this signaling protein modulates water channel function. Molecular dynamics and functional mutation studies reveal how CaM binding inhibits AQP0 water permeability by allosterically closing the cytoplasmic gate of AQP0. Our mechanistic model provides new insight, only possible in the context of the fully assembled channel, into how CaM regulates multimeric channels by facilitating cooperativity between adjacent subunits.

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Section: Discussionmentioning

confidence: 99%

Allosteric mechanism of water-channel gating by Ca2+–calmodulin

Reichow

Clemens

Freites

et al. 2013

Nat Struct Mol Biol

106

138

View full text Add to dashboard Cite

show abstract

“…As in SASREF a SA minimization is used to locate a best fitting arrangement of the rigid bodies and also the optimal local conformation of DRs. This approach has been used to successfully determine the structures of multi-domain proteins with flexible linkers (Clantin et al, 2010;Gut et al, 2009;Kozlov et al, 2009;NemethPongracz et al, 2007;Schmidt et al, 2010), and also for the addition of missing portions to crystal structures (Gut et al, 2009).…”

Section: Rigid Body Modelingmentioning

confidence: 99%

Structural characterization of proteins and complexes using small-angle X-ray solution scattering

Mertens

Svergun

2010

Journal of Structural Biology

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427

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“…The pK of the distal imidazolic nitrogen of His 465 affects the pH-dependent spectroscopic and catalytic properties of E. coli GadB, and His 465 must be present to integrate the cooperativity of triple helix formation into the cooperativity of the whole system. Notably, in the eukaryotic relative of E. coli GadB, Arabidopsis thaliana Gad1, this residue is not found in the long C-terminal tail, instrumental to activity regulation by pH and by Ca 2ϩ /calmodulin binding, and this probably explains why Gad1 does not undergo the same mechanism of inactivation (19).…”

mentioning

confidence: 99%

Mutation of His465 Alters the pH-dependent Spectroscopic Properties of Escherichia coli Glutamate Decarboxylase and Broadens the Range of Its Activity toward More Alkaline pH

Pennacchietti

Lammens

Capitani

et al. 2009

Journal of Biological Chemistry

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Glutamate decarboxylase (GadB) from Escherichia coli is a hexameric, pyridoxal 5-phosphate-dependent enzyme catalyzing CO 2 release from the ␣-carboxyl group of L-glutamate to yield ␥-aminobutyrate. GadB exhibits an acidic pH optimum and undergoes a spectroscopically detectable and strongly cooperative pH-dependent conformational change involving at least six protons. Crystallographic studies showed that at mildly alkaline pH GadB is inactive because all active sites are locked by the C termini and that the 340 nm absorbance is an aldamine formed by the pyridoxal 5-phosphate-Lys 276 Schiff base with the distal nitrogen of His 465 , the penultimate residue in the GadB sequence. Herein we show that His 465 has a massive influence on the equilibrium between active and inactive forms, the former being favored when this residue is absent. His 465 contributes with n ≈ 2.5 to the overall cooperativity of the system. The residual cooperativity (n ≈ 3) is associated with the conformational changes still occurring at the N-terminal ends regardless of the mutation. His 465 , dispensable for the cooperativity that affects enzyme activity, is essential to include the conformational change of the N termini into the cooperativity of the whole system. In the absence of His 465 , a 330-nm absorbing species appears, with fluorescence emission spectra more complex than model compounds and consisting of two maxima at 390 and 510 nm. Because His 465 mutants are active at pH well above 5.7, they appear to be suitable for biotechnological applications.

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A Common Structural Basis for pH- and Calmodulin-mediated Regulation in Plant Glutamate Decarboxylase

Cited by 76 publications

References 66 publications

Allosteric mechanism of water-channel gating by Ca2+–calmodulin

Allosteric mechanism of water-channel gating by Ca2+–calmodulin

Structural characterization of proteins and complexes using small-angle X-ray solution scattering

Mutation of His465 Alters the pH-dependent Spectroscopic Properties of Escherichia coli Glutamate Decarboxylase and Broadens the Range of Its Activity toward More Alkaline pH

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