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A fully atomistic model of the Cx32 connexon.

Pantano S, Zonta F, Mammano F - PLoS ONE (2008)

Bottom Line: The lack of high resolution data for connexon structures has hampered so far the study of the structure-function relationships that link molecular effects of disease-causing mutations with their observed phenotypes.Our results provide new mechanistic insights into the effects of numerous spontaneous mutations and their implication in connexin-related pathologies.This model constitutes a step forward towards a structurally detailed description of the gap junction architecture and provides a structural platform to plan new biochemical and biophysical experiments aimed at elucidating the structure of connexin channels and hemichannels.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur of Montevideo, Montevideo, Uruguay. spantano@pasteur.edu.uy

ABSTRACT
Connexins are plasma membrane proteins that associate in hexameric complexes to form channels named connexons. Two connexons in neighboring cells may dock to form a "gap junction" channel, i.e. an intercellular conduit that permits the direct exchange of solutes between the cytoplasm of adjacent cells and thus mediate cell-cell ion and metabolic signaling. The lack of high resolution data for connexon structures has hampered so far the study of the structure-function relationships that link molecular effects of disease-causing mutations with their observed phenotypes. Here we present a combination of modeling techniques and molecular dynamics (MD) to infer side chain positions starting from low resolution structures containing only C alpha atoms. We validated this procedure on the structure of the KcsA potassium channel, which is solved at atomic resolution. We then produced a fully atomistic model of a homotypic Cx32 connexon starting from a published model of the C alpha carbons arrangement for the connexin transmembrane helices, to which we added extracellular and cytoplasmic loops. To achieve structural relaxation within a realistic environment, we used MD simulations inserted in an explicit solvent-membrane context and we subsequently checked predictions of putative side chain positions and interactions in the Cx32 connexon against a vast body of experimental reports. Our results provide new mechanistic insights into the effects of numerous spontaneous mutations and their implication in connexin-related pathologies. This model constitutes a step forward towards a structurally detailed description of the gap junction architecture and provides a structural platform to plan new biochemical and biophysical experiments aimed at elucidating the structure of connexin channels and hemichannels.

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Inter connexin surface.Open-book representation of the average connexin dimer. The thick residues are those involved in protein–protein interactions reported in Table 3. Ribbons are colored by position going from red, to green and to blue from the center of the dimer.
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pone-0002614-g004: Inter connexin surface.Open-book representation of the average connexin dimer. The thick residues are those involved in protein–protein interactions reported in Table 3. Ribbons are colored by position going from red, to green and to blue from the center of the dimer.

Mentions: As shown in Figure 4, contacts between adjacent connexins, named A and B, are mediated by helices TM1 and TM3 in A, and by TM3 and TM4 in B. As previously observed, TM2 is the only helix that does not participate in inter−connexin contacts. In our simulations, these were essentially of hydrophobic nature and no firmly established electrostatic interactions were detected between adjacent connexins within the TM region. The average interface area between connexon subunits is 540 Å2, with 15 % and 7 % of polar atoms in the interfaces of connexin subunits A and B, respectively. Notably, several aromatic residues are located within the interface (two in connexin A and 5 in connexin B, Table 3), which could further strengthen the quaternary complex through aromatic−aromatic stacking interactions.


A fully atomistic model of the Cx32 connexon.

Pantano S, Zonta F, Mammano F - PLoS ONE (2008)

Inter connexin surface.Open-book representation of the average connexin dimer. The thick residues are those involved in protein–protein interactions reported in Table 3. Ribbons are colored by position going from red, to green and to blue from the center of the dimer.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2481295&req=5

pone-0002614-g004: Inter connexin surface.Open-book representation of the average connexin dimer. The thick residues are those involved in protein–protein interactions reported in Table 3. Ribbons are colored by position going from red, to green and to blue from the center of the dimer.
Mentions: As shown in Figure 4, contacts between adjacent connexins, named A and B, are mediated by helices TM1 and TM3 in A, and by TM3 and TM4 in B. As previously observed, TM2 is the only helix that does not participate in inter−connexin contacts. In our simulations, these were essentially of hydrophobic nature and no firmly established electrostatic interactions were detected between adjacent connexins within the TM region. The average interface area between connexon subunits is 540 Å2, with 15 % and 7 % of polar atoms in the interfaces of connexin subunits A and B, respectively. Notably, several aromatic residues are located within the interface (two in connexin A and 5 in connexin B, Table 3), which could further strengthen the quaternary complex through aromatic−aromatic stacking interactions.

Bottom Line: The lack of high resolution data for connexon structures has hampered so far the study of the structure-function relationships that link molecular effects of disease-causing mutations with their observed phenotypes.Our results provide new mechanistic insights into the effects of numerous spontaneous mutations and their implication in connexin-related pathologies.This model constitutes a step forward towards a structurally detailed description of the gap junction architecture and provides a structural platform to plan new biochemical and biophysical experiments aimed at elucidating the structure of connexin channels and hemichannels.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur of Montevideo, Montevideo, Uruguay. spantano@pasteur.edu.uy

ABSTRACT
Connexins are plasma membrane proteins that associate in hexameric complexes to form channels named connexons. Two connexons in neighboring cells may dock to form a "gap junction" channel, i.e. an intercellular conduit that permits the direct exchange of solutes between the cytoplasm of adjacent cells and thus mediate cell-cell ion and metabolic signaling. The lack of high resolution data for connexon structures has hampered so far the study of the structure-function relationships that link molecular effects of disease-causing mutations with their observed phenotypes. Here we present a combination of modeling techniques and molecular dynamics (MD) to infer side chain positions starting from low resolution structures containing only C alpha atoms. We validated this procedure on the structure of the KcsA potassium channel, which is solved at atomic resolution. We then produced a fully atomistic model of a homotypic Cx32 connexon starting from a published model of the C alpha carbons arrangement for the connexin transmembrane helices, to which we added extracellular and cytoplasmic loops. To achieve structural relaxation within a realistic environment, we used MD simulations inserted in an explicit solvent-membrane context and we subsequently checked predictions of putative side chain positions and interactions in the Cx32 connexon against a vast body of experimental reports. Our results provide new mechanistic insights into the effects of numerous spontaneous mutations and their implication in connexin-related pathologies. This model constitutes a step forward towards a structurally detailed description of the gap junction architecture and provides a structural platform to plan new biochemical and biophysical experiments aimed at elucidating the structure of connexin channels and hemichannels.

Show MeSH
Related in: MedlinePlus