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Liposomes, Exosomes, and Virosomes: From Modeling Complex
Membrane Processes to Medical Diagnostics and Drug Delivery
Poster Abstracts
95
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