Identificaiton of the pore-lining helices of connexin 43 gap-junctional hemichannels

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dc.contributor.advisor Guillermo A . Altenberg en_US
dc.contributor.committeeMember Robert O . Fox en_US
dc.contributor.committeeMember Krishna Rajarathnam en_US
dc.contributor.committeeMember Juan Carlos Sáez en_US
dc.contributor.committeeMember Andrés Oberhauser en_US
dc.creator Sung -Chang Lee en_US
dc.date.accessioned 2011 -12 -20T16 :04 :33Z
dc.date.accessioned 2014 -02 -19T22 :05 :02Z
dc.date.available 2008 -06 -17 en_US
dc.date.available 2011 -12 -20T16 :04 :33Z
dc.date.available 2014 -02 -19T22 :05 :02Z
dc.date.created 2008 -03 -31 en_US
dc.date.issued 2008 -03 -28 en_US
dc.identifier.other etd -03312008 -160050 en_US
dc.identifier.uri http : / /hdl .handle .net /2152 .3 /82
dc.description.abstract The gap -junctional channels that mediate intercellular communication are formed by head -to -head docking of two gap -junctional hemichannels from adjacent cells . The hemichannels are hexamers of connexins , proteins that have four transmembrane helices . The transmembrane helices that line the gap -junctional pore have not been identified , and their identification was the main goal of my dissertation project . To accomplish this goal , I used a combination of molecular biology , biochemical and biophysical techniques that include poly -alanine helix scanning mutagenesis , the substituted cysteine accessibility method and luminescence resonance energy transfer . Using the latter methodology in particular , as well as a new method to produce purified hemichannels of controlled subunit composition , I was able to assign all helices in the available low -resolution cryoelectron microscopy structure published by others , where helices are named A through D , and generate the first model of gap -junctional channels and hemichannels based on experimental structural measurements . In this model , connexin transmembrane helices 1 through 4 correspond to helices A , C , B and D , respectively . Luminescence resonance energy transfer is a powerful method for structural studies of membrane proteins in their native bilayer environment . Taken advantage of this methodology , in combination with the generation of hemichannels of controlled subunit composition , I was also able to determine that PKC -mediated phosphorylation of Ser368 produces a partial closure of the Cx43 hemichannel pore , that this effects requires phosphorylation of all six Cx43 monomers in the hemichannel , and that the decrease in permeability is accompanied by significant conformational changes of the connexin molecules . These changes involve increases of the distances separating the C -terminal ends of the subunits and decreases in the distances separating the pore -lining helices ; both changes in inter -subunit distances are of the order of several Angstroms . These results indicate that a simple ball -and -chain mechanism cannot explain the gating of Cx43 hemichannels by PKC -mediated phosphorylation and that a significant re -arrangement of pore helices takes place instead . In summary , my results allowed me to generate an experimentally -based model gap -junctional channels and hemichannels and to gain insight into the molecular mechanism of Cx43 regulation by PKC -mediated phosphorylation . en_US
dc.format.medium electronic en_US
dc.language.iso eng en_US
dc.rights Copyright © is held by the author . Presentation of this material on the TDL web site by The University of Texas Medical Branch at Galveston was made possible under a limited license grant from the author who has retained all copyrights in the works . en_US
dc.subject gap junctional channel structure en_US
dc.subject Connexin 43 en_US
dc.title Identificaiton of the pore -lining helices of connexin 43 gap -junctional hemichannels en_US
dc.type.genre dissertation en_US
dc.type.material text en_US
thesis.degree.name PhD en_US
thesis.degree.level Doctoral en_US
thesis.degree.grantor The University of Texas Medical Branch en_US
thesis.degree.department Cellular Physiology and Molecular Biophysics en_US

Citation

Identificaiton of the pore-lining helices of connexin 43 gap-junctional hemichannels. Doctoral dissertation, The University of Texas Medical Branch. Available electronically from http : / /hdl .handle .net /2152 .3 /82 .

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03introduction.pdf 230.9Kb application/pdf View/Open
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