Please use this identifier to cite or link to this item: doi:10.22028/D291-29238
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Title: The stoichiometry of the TMEM16A ion channel determined in intact plasma membranes of COS-7 cells using liquid-phase electron microscopy
Author(s): Peckys, Diana B.
Stoerger, Christof
Latta, Lorenz
Wissenbach, Ulrich
Flockerzi, Veit
de Jonge, Niels
Language: English
Title: Journal of structural biology
Volume: 199
Issue: 2
Startpage: 102
Endpage: 113
Pages: 102-113
Publisher/Platform: Elsevier
Year of Publication: 2017
Publikation type: Journal Article
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 of the first publication: 10.1016/j.jsb.2017.05.009
Link to this record: hdl:20.500.11880/28046
ISSN: 1047-8477
Date of registration: 10-Oct-2019
Faculty: NT - Naturwissenschaftlich- Technische Fakultät
Department: NT - Physik
Professorship: NT - Keiner Professur zugeordnet
Collections:Die Universitätsbibliographie

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