title: Conformational rearrangements in the transmembrane domain of CNGA1 channels revealed by single-molecule force spectroscopy.
abstract: Cyclic nucleotide-gated (CNG) channels are activated by binding of cyclic nucleotides. Although structural studies have identified the channel pore and selectivity filter, conformation changes associated with gating remain poorly understood. Here we combine single-molecule force spectroscopy (SMFS) with mutagenesis, bioinformatics and electrophysiology to study conformational changes associated with gating. By expressing functional channels with SMFS fingerprints in Xenopus laevis oocytes, we were able to investigate gating of CNGA1 in a physiological-like membrane. Force spectra determined that the S4 transmembrane domain is mechanically coupled to S5 in the closed state, but S3 in the open state. We also show there are multiple pathways for the unfolding of the transmembrane domains, probably caused by a different degree of α-helix folding. This approach demonstrates that CNG transmembrane domains have dynamic structure and establishes SMFS as a tool for probing conformational change in ion channels.
authors.affiliation: International School for Advanced Studies (SISSA) Neuroscience Area, via Bonomea 265, Trieste 34136, Italy.
authors.forename: Sourav
authors.initials: S
authors.lastname: Maity
authors.affiliation: International School for Advanced Studies (SISSA) Neuroscience Area, via Bonomea 265, Trieste 34136, Italy.|CBM S.c.r.l., Area Science Park, Basovizza, Trieste 34149, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Mazzolini
authors.affiliation: International School for Advanced Studies (SISSA) Neuroscience Area, via Bonomea 265, Trieste 34136, Italy.
authors.forename: Manuel
authors.initials: M
authors.lastname: Arcangeletti
authors.affiliation: International School for Advanced Studies (SISSA) Neuroscience Area, via Bonomea 265, Trieste 34136, Italy.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Valbuena
authors.affiliation: International School for Advanced Studies (SISSA) Neuroscience Area, via Bonomea 265, Trieste 34136, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Fabris
authors.affiliation: CBM S.c.r.l., Area Science Park, Basovizza, Trieste 34149, Italy.|IOM-CNR, Area Science Park, Basovizza, Trieste 34149, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Lazzarino
authors.affiliation: International School for Advanced Studies (SISSA) Neuroscience Area, via Bonomea 265, Trieste 34136, Italy.
authors.forename: Vincent
authors.initials: V
authors.lastname: Torre
mesh_terms: D000818:Animals; D019295:Computational Biology; D054815:Cyclic Nucleotide-Gated Cation Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D055724:Electrophysiological Phenomena; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D015640:Ion Channel Gating / Q000502:physiology*; D006080:Ovarian Follicle; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013057:Spectrum Analysis / Q000379:methods*; D014982:Xenopus laevis
chemical_list: C475741:CNGA1 protein, bovine; D054815:Cyclic Nucleotide-Gated Cation Channels