PUBLICATION
Molecular dynamics and in vitro analysis of Connexin43: A new 14-3-3 mode-1 interacting protein
- Authors
- Park, D.J., Freitas, T.A., Wallick, C.J., Guyette, C.V., and Warn-Cramer, B.J.
- ID
- ZDB-PUB-061010-12
- Date
- 2006
- Source
- Protein science : a publication of the Protein Society 15(10): 2344-2355 (Journal)
- Registered Authors
- Keywords
- Connexin43, 14-3-3, protein?protein interactions, homology modeling, phosphorylation, mode-1 binding motif
- MeSH Terms
-
- Rats
- Binding Sites
- Phosphorylation
- Connexin 43/genetics
- Connexin 43/metabolism*
- Connexin 43/physiology
- Multiprotein Complexes
- Animals
- Cell Line
- Proto-Oncogene Proteins c-akt/antagonists & inhibitors
- Cell Communication
- Species Specificity
- Amino Acid Substitution
- 14-3-3 Proteins/metabolism*
- Protein Binding
- Serine/metabolism
- PubMed
- 17008717 Full text @ Protein Sci.
Citation
Park, D.J., Freitas, T.A., Wallick, C.J., Guyette, C.V., and Warn-Cramer, B.J. (2006) Molecular dynamics and in vitro analysis of Connexin43: A new 14-3-3 mode-1 interacting protein. Protein science : a publication of the Protein Society. 15(10):2344-2355.
Abstract
The interaction of cellular proteins with the gap junction protein Connexin43 (Cx43) is thought to form a dynamic scaffolding complex that functions as a platform for the assembly of signaling, structural, and cytoskeletal proteins. A high stringency Scansite search of rat Cx43 identified the motif containing Ser373 (S373) as a 14-3-3 binding site. The S373 motif and the second best mode-1 motif, containing Ser244 (S244), are conserved in rat, mouse, human, chicken, and bovine, but not in Xenopus or zebrafish Cx43. Docking studies of a mouse/rat 14-3-3 homology model with the modeled phosphorylated S373 or S244 peptide ligands or their serine-to-alanine mutants, S373A or S244A, revealed that the pS373 motif facilitated a greater number of intermolecular contacts than the pS244 motif, thus supporting a stronger 14-3-3 binding interaction with the pS373 motif. The alanine substitution also reduced more than half the number of intermolecular contacts between 14-3-3 and the S373 motif, emphasizing the phosphorylation dependence of this interaction. Furthermore, the ability of the wild-type or the S244A GST-Cx43 C-terminal fusion protein, but not the S373A fusion protein, to interact with either 14-3-3 or 14-3-3zeta in GST pull-down experiments clearly demonstrated that the S373 motif mediates the direct interaction between Cx43 and 14-3-3 proteins. Blocking growth factor-induced Akt activation and presumably any Akt-mediated phosphorylation of the S373 motif in ROSE 199 cells did not prevent the down-regulation of Cx43-mediated cell-cell communication, suggesting that an Akt-mediated interaction with 14-3-3 was not involved in the disruption of Cx43 function.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping