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Related Concept Videos

Calmodulin-dependent Signaling01:16

Calmodulin-dependent Signaling

Calmodulin (CaM) is a calcium-binding protein in eukaryotes that controls various calcium-regulated cellular processes. It has four calcium-binding sites that bind calcium to form the calcium-calmodulin ( Ca2+-CaM) complex. GPCR stimulation increases the calcium levels in the cells that bind to CaM and induces a conformational change.
The Ca2+-CaM complex does not have enzymatic activity by itself. Instead, the complex binds downstream target proteins, including membrane proteins or enzymes,...
Cooperative Allosteric Transitions01:58

Cooperative Allosteric Transitions

Cooperative allosteric transitions can occur in multimeric proteins, where each subunit of the protein has its own ligand-binding site. When a ligand binds to any of these subunits, it triggers a conformational change that affects the binding sites in the other subunits; this can change the affinity of the other sites for their respective ligands. The ability of the protein to change the shape of its binding site is attributed to the presence of a mix of flexible and stable segments in the...
Cooperative Allosteric Transitions01:58

Cooperative Allosteric Transitions

Cooperative allosteric transitions can occur in multimeric proteins, where each subunit of the protein has its own ligand-binding site. When a ligand binds to any of these subunits, it triggers a conformational change that affects the binding sites in the other subunits; this can change the affinity of the other sites for their respective ligands. The ability of the protein to change the shape of its binding site is attributed to the presence of a mix of flexible and stable segments in the...
Cooperative Allosteric Transitions01:58

Cooperative Allosteric Transitions

Cooperative allosteric transitions can occur in multimeric proteins, where each subunit of the protein has its own ligand-binding site. When a ligand binds to any of these subunits, it triggers a conformational change that affects the binding sites in the other subunits; this can change the affinity of the other sites for their respective ligands. The ability of the protein to change the shape of its binding site is attributed to the presence of a mix of flexible and stable segments in the...
Ligand-gated Ion Channels01:19

Ligand-gated Ion Channels

Ligand-gated ion channels are transmembrane proteins with a channel for ions to pass through and a binding site for a ligand. The channel opens only when a ligand attaches to the binding site.
Three Subfamilies of Ligand-gated Ion Channels
Ligand-gated ion channels fall into three subfamilies. The 'Cys-loop' includes the nicotinic acetylcholine receptors, γ-aminobutyric acid (GABA), glycine, and 5-hydroxytryptamine receptors. The second one is the 'Pore-loop' channels that include the...
Ligand-gated Ion Channels01:19

Ligand-gated Ion Channels

Ligand-gated ion channels are transmembrane proteins with a channel for ions to pass through and a binding site for a ligand. The channel opens only when a ligand attaches to the binding site.
Three Subfamilies of Ligand-gated Ion Channels
Ligand-gated ion channels fall into three subfamilies. The 'Cys-loop' includes the nicotinic acetylcholine receptors, γ-aminobutyric acid (GABA), glycine, and 5-hydroxytryptamine receptors. The second one is the 'Pore-loop' channels that include the...

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Pull-down of Calmodulin-binding Proteins
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Published on: January 23, 2012

Cooperativity between calmodulin-binding sites in Kv7.2 channels.

Alessandro Alaimo1, Araitz Alberdi, Carolina Gomis-Perez

  • 1Unidad de Biofísica, CSIC-UPV/EHU, Universidad del País Vasco, Barrio Sarriena s/n, 48940 Leioa, Spain.

Journal of Cell Science
|December 4, 2012
PubMed
Summary

Calmodulin (CaM) binding to Kv7.2 channels is essential for their ER exit and surface expression. This CaM interaction influences channel function and is linked to benign familial neonatal convulsions.

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Area of Science:

  • Neuroscience
  • Molecular Biology
  • Biochemistry

Background:

  • Calmodulin (CaM) uniquely controls Kv7.2 channel surface expression via two binding sites.
  • Impaired CaM binding or sequestration halts Kv7.2 channel ER exit, reducing M-current and increasing neuronal excitability.
  • This mechanism offers insight into benign familial neonatal convulsions (BFNC).

Purpose of the Study:

  • To investigate the role of CaM binding in Kv7.2 channel trafficking and function.
  • To re-evaluate CaM binding to a Kv7.2 channel mutant (S511D) previously thought to bypass CaM dependency.
  • To elucidate the binding affinity and cooperative interaction of CaM with Kv7.2 channel's binding domains.

Main Methods:

  • Biochemical techniques to detect CaM binding to wild-type and S511D mutant Kv7.2 channels.
  • Functional assays in HEK293 cells to assess surface expression and rescue of channel function.
  • Affinity measurements for CaM binding to helix A and helix B within the CaM-binding domain.

Main Results:

  • The S511D mutant possesses functional CaM-binding sites, previously undetected by standard assays.
  • CaM rescues surface expression and function of the S511D mutant, indicating limiting free CaM in cells.
  • CaM exhibits higher affinity for helix B than helix A, with cooperative binding between the two sites.
  • CaM can bridge subunits, potentially influencing Kv7.2 channel function through inter-subunit interactions.

Conclusions:

  • CaM binding is required for Kv7.2 channel ER exit and subsequent surface expression.
  • The S511D mutation does not abolish CaM binding but alters its detectability.
  • CaM's cooperative binding to Kv7.2 channels, involving distinct affinities for helix A and B, is crucial for channel regulation.