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Titel: The stoichiometry of the TMEM16A ion channel determined in intact plasma membranes of COS-7 cells using liquid-phase electron microscopy
VerfasserIn: Peckys, Diana B.
Stoerger, Christof
Latta, Lorenz
Wissenbach, Ulrich
Flockerzi, Veit
de Jonge, Niels
Sprache: Englisch
Titel: Journal of structural biology
Bandnummer: 199
Heft: 2
Startseite: 102
Endseite: 113
Seiten: 102-113
Verlag/Plattform: Elsevier
Erscheinungsjahr: 2017
Dokumenttyp: Journalartikel / Zeitschriftenartikel
Abstract: TMEM16A is a membrane protein forming a calcium-activated chloride channel. A homodimeric stoichiometry of the TMEM16 family of proteins has been reported but an important question is whether the protein resides always in a dimeric configuration in the plasma membrane or whether monomers of the protein are also present in its native state within in the intact plasma membrane. We have determined the stoichiometry of the human (h)TMEM16A within whole COS-7 cells in liquid. For the purpose of detecting TMEM16A subunits, single proteins were tagged by the streptavidin-binding peptide within extracellular loops accessible by streptavidin coated quantum dot (QD) nanoparticles. The labeled proteins were then imaged using correlative light microscopy and environmental scanning electron microscopy (ESEM) using scanning transmission electron microscopy (STEM) detection. The locations of 19,583 individual proteins were determined of which a statistical analysis using the pair correlation function revealed the presence of a dimeric conformation of the protein. The amounts of detected label pairs and single labels were compared between experiments in which the TMEM16A SBP-tag position was varied, and experiments in which tagged and non-tagged TMEM16A proteins were present. It followed that hTMEM16A resides in the plasma membrane as dimer only and is not present as monomer. This strategy may help to elucidate the stoichiometry of other membrane protein species within the context of the intact plasma membrane in future.
DOI der Erstveröffentlichung: 10.1016/j.jsb.2017.05.009
Link zu diesem Datensatz: hdl:20.500.11880/28046
http://dx.doi.org/10.22028/D291-29238
ISSN: 1047-8477
Datum des Eintrags: 10-Okt-2019
Fakultät: NT - Naturwissenschaftlich- Technische Fakultät
Fachrichtung: NT - Physik
Professur: NT - Keiner Professur zugeordnet
Sammlung:SciDok - Der Wissenschaftsserver der Universität des Saarlandes

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