Biophysical Society Thematic Meeting | Ascona, Switzerland

Liposomes, Exosomes, and Virosomes: From Modeling Complex Membrane Processes to Medical Diagnostics and Drug Delivery

Poster Abstracts

3-POS Board 2 Functionalized SMA-Nanodiscs for Conjugation of Membrane Proteins to Dyes and Surfaces Simon Lindhoud, Vanessa Carvalho, Jochem Pronk, Marie-Eve Aubin-Tam . Delft University of Technology, Delft, Netherlands. The integration of membrane proteins into biophysical and biosensing assays have lagged considerably in comparison to soluble proteins owing to the extreme difficulties associated with purifying, handling and conjugating membrane proteins. I will present newly developed functionalization procedures to interface membrane proteins with surfaces, while preserving their native membrane environment. Nanodiscs are soluble scaffolds for membrane proteins, which consist of nanometer-sized discoidal phospholipid bilayers surrounded by an amphipatic polymer, such as the membrane scaffold protein [1] or styrene-maleic acid (SMA) co-polymer [2]. Membrane proteins embedded in lipid nanodiscs maintain their membrane-integrated state in this soluble complex, and can therefore be handled similarly to soluble proteins. SMA co-polymers are highlighted as agents for detergent-free purification of membrane proteins, which can solubilize membrane proteins in presence of their native lipid membrane environment. To facilitate conjugation of SMA- nanodiscs to surfaces, nanoparticles and fluorophores, we exploit the reactivity of maleic anhydride moieties in SMA towards amines to modify the polymer with cysteamine [3]. Thus, we equip the polymer with a sulfhydril group (SMA-SH). This sulfhydryl group is then modified with thiol-reactive probes, such as maleimide derivatives of fluorophores and biotin. SMA-SH nanodiscs are characterized with gel filtration, dynamic light scattering and electron microscopy. We find that SMA-SH enables the functionalization of membrane proteins with a variety of different probes, while not requiring any mutation or chemical modification of the protein itself. We anticipate that this versatile approach will find application in membrane protein purification, in biosensing, and in biophysical assays. 1. Bayburt TH, Grinkova YV, Sligar SG. Nano Lett., 2, 853-856, 2002 2. Scheidelaar, S., et al. Biophys. J., 108, 279-290, 2015 3. Lindhoud S, Carvalho V, Pronk JW, Aubin-Tam ME, Biomacromolecules, 17, 1516-1522, 2016

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