Document text
Principal Investigator: Weihua Qiu
Organization: VIRGINIA COMMONWEALTH UNIVERSITY
Fiscal Year: 2021
Award: $229,008
Funding agency: National Institute on Deafness and Other Communication Disorders
SUMMARY
Connexin 26, as a gap junction channel, directly regulates the cell-to-cell exchange of ion,
metabolites, and second messengers. Mutations of connexin 26 can cause severe hearing loss.
However, the molecular gating mechanism on the structural basis of connexin 26 is still unclear,
especially the closed state and malfunction of connexin 26 mutations. We have developed the
native cell membrane nanoparticle (NCMN) system for high-resolution single-particle cryo-EM
structure determination and functional studies of membrane proteins within their physiological
conditions. We recently solved the cryo-EM structures of connexin 26 in the absence or presence
of calcium ions in an open state at 2.3 Å and 2.9 Å associated with about 36 lipid molecules via
the NCMN system. However, the close state of connexin 26 with high resolution is still uncovered.
We hypothesize that our NCMN system's application to structure determination of Cx26 and
Cx26-R75W in the presence and absence of calcium and calmodulin can provide unique
structural information. Aim 1: To investigate the CLOSED conformation of the gating mechanism
of Cx26. Aim 2: To elucidate the molecular basis of how the single point mutation R75W in Cx26
leads to profound hearing loss. The proposed experimental results will provide insights into
detailed structural information and biochemical analysis in understanding the gating mechanism,
also provide mechanistic details on abnormal channel gating function detected in this hearing
disorder. The structural information may lead to some novel strategy to modulate the malfunction
CX26-R75W for hearing loss.
Terms: <Amino Acids><Assay><Bioassay><Biochemical><Biologic Assays><Biological Assay><CX26><Calcium><Calcium ion><Calcium-Dependent Activator Protein><Calcium-Dependent Regulator><Calmodulin><Cell Body><Cell Function><Cell Process><Cell membrane><Cell physiology><Cell to Cell Communication and Signaling><Cell-Cell Signaling><Cells><Cellular Function><Cellular Physiology><Cellular Process><Chemicals><Clinical><Coloring Agents><Complex><Connexin-26><Connexins><Cryo-electron Microscopy><Cryoelectron Microscopy><Cytoplasmic Membrane><Data><Detergents><Dyes><Dysfunction><Electron Cryomicroscopy><Environment><Family><Functional disorder><GJB2><GJB2 gene><Gap Junction Protein, 26-KD><Gap Junction Protein, Beta-2><Gap Junction Proteins><Genetic Alteration><Genetic Change><Genetic defect><Glean><HeLa><Hearing Disorders><Hearing Loss><Hearing problem><Hela Cells><Human><Human Figure><Human body><Hypoacuses><Hypoacusis><Intracellular Second Messenger><Ion Exchange><KID syndrome><Lead><Lipids><Membrane><Membrane Protein Gene><Membrane Proteins><Membrane-Associated Proteins><Modern Man><Molecular><Molecular Configuration><Molecular Conformation><Molecular Stereochemistry><Mutation><Organism-Level Process><Organismal Process><Outcome><Pathology><Pb element><Phosphodiesterase Activating Factor><Phosphodiesterase Protein Activator><Physiologic><Physiologic Processes><Physiological><Physiological Processes><Physiopathology><Plasma Membrane><Play><Point Mutation><Polymers><Proteins><Protocol><Protocols documentation><Resolution><Role><Sampling><Second Messenger Systems><Second Messengers><Senter syndrome><Solid><Structure><Subcellular Process><Surface Proteins><System><TM Domain><Testing><Transmembrane Domain><Transmembrane Region><aminoacid><auditory disease><auditory disorder><auditory dysfunction><auditory problem><base><conformation><conformational state><cryo-EM><cryoEM><deaf><deafened><density><dysfunctional hearing><gap junction channel><genome mutation><hearing defect><hearing deficit><hearing difficulty><hearing disability><hearing disease><hearing dysfunction><hearing impairment><heavy metal Pb><heavy metal lead><improved><insight><intercellular communication><interest><keratitis-ichthyosis-deafness syndrome><member><membrane structure><mutant><nano particle><nano-sized particle><nanoparticle><nanosized particle><native protein drug><new approaches><novel approaches><novel strategies><novel strategy><particle><pathophysiology><pharmaceutical protein><plasmalemma><profound hearing loss><protein complex><protein drug agent><signal transduction second messengers><small molecule><social role><therapeutic protein>