Structure-Function relation of Connexin disease mutations


Principal Investigator: TERRY L DOWD
Abstract: DESCRIPTION (provided by applicant): The N-terminal domain (NT, residues 1-22) is an important determinant of perm-selectivity and voltage-dependent gating of connexin channels and a sensitive mutational target underlying two common inherited diseases: X-linked Charcot-Marie-Tooth (Cx32) and nonsyndromic and syndromic deafness (Cx26). This proposal will determine how disease causing mutations in the NT of Cx32 and Cx26 alter channel function and channel biosynthesis by applying synergistic computational and experimental approaches. Differences in function between wild type and disease causing NT mutations are hypothesized to arise from specific changes in channel structure. The study will examine 9 NT loci comprising mutations in both Cx32 and Cx26. In several cases, mutations of the same locus alter Cx26 and Cx32 channel function differently, suggesting that identical or homologous amino acid substitutions cause different structural defects in the two connexins. Studies will be guided by the crystal structure of a Cx26 hemichannel and a Cx32 homology model, both refined by all-atom molecular dynamics (MD) simulation and shown to closely correspond to the structure of the biological open channel. The study will solve the structure of mutant NT peptides by 2D NMR. Structural solutions of longer wild-type and mutant peptides (NT-CL domain, residues 1-114) in a membrane environment by 3D NMR, and assembled channels by x-ray crystallography have been initiated. Resulting atomic models of connexin channels will be refined by all-atom MD simulations, the permeabilities to ions and second messengers determined computationally and compared to experimental. This experimental strategy provides a sensitive test of the accuracy of atomic models, insights into molecular mechanisms of perm-selectivity and how these are changed by mutation, as well as testable hypotheses of structure-function relations. The study will investigate the role of the NT in channel biogenesis by determining the position and stability of the NT of connexin subunits inserted into canine microsomal membranes, the role of the NT in subunit oligomerization, and when and how the NT assumes its final position deep within the pore of assembled hemichannels prior to plasma membrane insertion. Parallel computational studies will provide a rigorous mechanistic framework that will guide these experimental studies. This new, fundamental knowledge will provide a framework for understanding the molecular defects of the class of disease causing NT mutations that are not plasma membrane inserted but trapped in cytosolic compartments and targeted for degradation. The project is highly collaborative, bringing together investigators with proven expertise in structural determination, computational methods and biophysical characterization of connexin channels. The results will provide new information fundamental to the elucidation of connexin disease etiology and to the development of effective treatments.
Funding Period: 2012-09-01 - 2016-08-31
more information: NIH RePORT

Top Publications

  1. pmc The carboxyl terminal residues 220-283 are not required for voltage gating of a chimeric connexin32 hemichannel
    Taekyung Kwon
    Dominic P Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York
    Biophys J 105:1376-82. 2013

Research Grants

Detail Information


  1. pmc The carboxyl terminal residues 220-283 are not required for voltage gating of a chimeric connexin32 hemichannel
    Taekyung Kwon
    Dominic P Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York
    Biophys J 105:1376-82. 2013
    ..We conclude that an operational Cx32 Vj (fast) gate does not require CT residues 220-283, as reported previously by others. ..

Research Grants30

  1. The Transmembrane Protein Center
    Brian G Fox; Fiscal Year: 2013
    ..Expanded options for crystallization screening, access to synchrotrons, and improvements in software used to solve structures will also contribute to the increased throughput necessary to achieve PSI:Biology goals. ..
  2. Genomics of Kidney Transplantation Admin
    Arthur J Matas; Fiscal Year: 2013
    ..The Administrative Core will also facilitate interaction between Sites, Cores, and Projects. ..
  3. Structural and Computational Studies of LOV domain proteins and Cryptochromes
    Karen S Conrad; Fiscal Year: 2013
  4. Transplant Tolerance in Non-Human Primates
    ..This goal will be accomplished via four interrelated projects and two supporting cores. ..
  5. Molecular and Cellular Mechanisms of Vascular Anomalies
    Bjorn Reino Olsen; Fiscal Year: 2013
  6. Center for Structure of Membrane Proteins
    Robert M Stroud; Fiscal Year: 2013
    ..3.1, one of the world's most productive protein crystallography facilities. Overall, the combined expertise of principal investigators provides a unique environment to achieve the proposed aims. ..
  7. Interdisciplinary center of excellence for the study of pain and sensory function
    Ian D Meng; Fiscal Year: 2013
    ..Completion of these Aims will develop the research careers of a multidisciplinary group of junior investigators, and establish the core facilities and equipment necessary to constitute a competitive research center. ..
  8. Structural Genomics on Membrane Proteins
    Wayne A Hendrickson; Fiscal Year: 2013
    ..The project will be managed to optimize output and to integrate effectively with the PSI-Biology network and with other membrane protein structure efforts. ..
  9. Innate/Adaptive Immune Interactions in Gut Inflammation
    Sergio A Lira; Fiscal Year: 2013
    ..Overall the focus and interactive nature of the program (20 joint publications in 4 years, 3 additional NIH grants on related areas) provide a solid basis for a productive outcome. ..
  10. Endothelial Cell Phenotypes in Health and Disease
    William C Aird; Fiscal Year: 2013
    ..Core C ("Gene Targeting Core";William C. Aird, Core Leader) provides the necessary tools for targeting the Hprt locus and the loci of endogenous genes in ES cells and mice. ..
    VYTAUTAS K VERSELIS; Fiscal Year: 2013
    ..These studies should also shed light on a growing list of disorders ascribed to hemichannel dysfunction that includes atherosclerosis, stroke, neuropathy and congenital cataractogenesis and should lead to strategies for treatment. ..
  12. Endogenous Cannabinoids and Brain Function
    Aron H Lichtman; Fiscal Year: 2013
    ..Ultimately, the knowledge gained from this basic research will yield novel therapeutic targets that can be exploited with the pharmacological agents developed here. PROGRAM CHARACTERISTICS ..
  13. Fundamental Studies of RNA Folding
    Daniel Herschlag; Fiscal Year: 2013
    ..abstract_text> ..
  14. Pacific NorthWest Regional Center of Excellence (PNWRCE)
    Jay A Nelson; Fiscal Year: 2013
    ..pseudomallei host pathogen response during both the septicemic as well as the intracellular phases of the disease. ..
    Gina E Sosinsky; Fiscal Year: 2013
    ..The proposed research is significant because results will be useful in defining better drugs and other therapeutics that potentially ameliorate connexin-related diseases. ..
  16. Cx26 mutations in syndromic deafness linked to skin disease
    Thomas W White; Fiscal Year: 2013
  17. Northeast Biodefense Center
    W Ian Lipkin; Fiscal Year: 2013
    ..As a Center based in a School of Public Health and a State Department of Health, the NBC has a firm commitment to and practical understanding of Emergency Preparedness. ..
  18. IkB/NF-kB Recognition In Silico, In Vitro and In Vivo
    Elizabeth A Komives; Fiscal Year: 2013
  19. Pacific Southwest RCE for Biodefense &Emerging Infectious Diseases Research
    Alan G Barbour; Fiscal Year: 2013
    ..abstract_text> ..
  20. Cyclic nucleotide permeability of connexin mutants related to genetic disease
    Virginijus Valiunas; Fiscal Year: 2013
    ..The results of the proposed research will establish a baseline for understanding the role of perm-selectivity as a potential determinant of disease states such as oculodentodigital dysplasia and atrial fibrillation. ..
  21. Rocky Mountain Regional Center of Excellence or Biodefense and Emerging Infectiou
    John T Belisle; Fiscal Year: 2013
    ..abstract_text> ..
    Kenneth H Cowan; Fiscal Year: 2013
  23. Gating and Regulation of Connexin Hemichannels
    JORGE ENRIQUE CONTRERAS; Fiscal Year: 2013
  24. Molecular motors in cell biology
    Yale E Goldman; Fiscal Year: 2013
    ..We anticipate that the proposed work will take us significantly further toward our goal of understanding motility in the normal and pathological function of cells. ..