A Three-Dimensional Model of the Yeast Transmembrane Sensor Wsc1 Obtained by SMA-Based Detergent-Free Purification and Transmission Electron Microscopy

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Title: A Three-Dimensional Model of the Yeast Transmembrane Sensor Wsc1 Obtained by SMA-Based Detergent-Free Purification and Transmission Electron Microscopy
Authors: Voskoboynikova, Natalia
Karlova, Maria
Kurre, Rainer
Mulkidjanian, Armen Y.
Shaitan, Konstantin V.
Sokolova, Olga S.
Steinhoff, Heinz-Jürgen
Heinisch, Jürgen J.
ORCID of the author: https://orcid.org/0000-0003-4197-4285
https://orcid.org/0000-0003-2317-0144
https://orcid.org/0000-0002-5137-303X
https://orcid.org/0000-0003-4678-232X
https://orcid.org/0000-0002-5888-0157
Abstract: The cell wall sensor Wsc1 belongs to a small family of transmembrane proteins, which are crucial to sustain cell integrity in yeast and other fungi. Wsc1 acts as a mechanosensor of the cell wall integrity (CWI) signal transduction pathway which responds to external stresses. Here we report on the purification of Wsc1 by its trapping in water-soluble polymer-stabilized lipid nanoparticles, obtained with an amphipathic styrene-maleic acid (SMA) copolymer. The latter was employed to transfer tagged sensors from their native yeast membranes into SMA/lipid particles (SMALPs), which allows their purification in a functional state, i.e., avoiding denaturation. The SMALPs composition was characterized by fluorescence correlation spectroscopy, followed by two-dimensional image acquisition from single particle transmission electron microscopy to build a three-dimensional model of the sensor. The latter confirms that Wsc1 consists of a large extracellular domain connected to a smaller intracellular part by a single transmembrane domain, which is embedded within the hydrophobic moiety of the lipid bilayer. The successful extraction of a sensor from the yeast plasma membrane by a detergent-free procedure into a native-like membrane environment provides new prospects for in vitro structural and functional studies of yeast plasma proteins which are likely to be applicable to other fungi, including plant and human pathogens.
Citations: Voskoboynikova N, Karlova M, Kurre R, Mulkidjanian AY, Shaitan KV, Sokolova OS, Steinhoff H-J, Heinisch JJ.: A Three-Dimensional Model of the Yeast Transmembrane Sensor Wsc1 Obtained by SMA-Based Detergent-Free Purification and Transmission Electron Microscopy. Journal of Fungi. 2021; 7(2):118.
URL: https://repositorium.ub.uni-osnabrueck.de/handle/urn:nbn:de:gbv:700-202106105037
Subject Keywords: Wsc1; membrane sensor; SMALP; detergent-free extraction; fluorescence correlation spectroscopy; transmission electron microscopy; 3D reconstruction
Issue Date: 5-Feb-2021
License name: Attribution 4.0 International
License url: http://creativecommons.org/licenses/by/4.0/
Type of publication: Einzelbeitrag in einer wissenschaftlichen Zeitschrift [article]
Appears in Collections:FB05 - Hochschulschriften
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