Giant plasma membrane vesicles (GPMVs) are large, unilamellar bodies that retain the exact membrane composition of the cells they are released from. In this work, we have isolated GPMVs from Human Embryonic Kidney (HEK) cells overexpressing an N-terminally SNAP-tagged G protein-coupled adenosine receptor. SNAP-tagged proteins rapidly react with benzylguanine derivatives to form an irreversible covalent bond. Thus, we generated a novel benzylguanine-PEG layer using chlorosilane and thiol-ene chemistry on glass for use in anchoring and optical monitoring of fluorescent GMPVs. Surface modification steps were verified using water contact angle and ellipsometry. Generation and characterization of fusion proteins and optical monitoring of GPMV-surface interactions are discussed herein. This system has the potential to be highly versatile, as potentially any protein of interest can be overexpressed in mammalian cells and selectively anchored in GPMVs using our method. This will greatly aid in a variety of downstream analyses, including reagentless optical biosensing.

Design and characterization of a benzylguanine-PEG surface for anchoring cellular-derived giant plasma membrane vesicles

255th National Meeting (2018)

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