Lipid bilayers on polyacrylamide brushes for inclusion of membrane proteins

Emily A. Smith, Jason W. Coym, Scott M. Cowell, Takahira Tokimoto, Victor J. Hruby, Henry I. Yamamura, Mary J. Wirth

Research output: Contribution to journalArticle

43 Scopus citations

Abstract

The ability of neutral polymer cushions to support neutral lipid bilayers for the incorporation of mobile transmembrane proteins was investigated. Polyacrylamide brush layers were grown on fused silica using atom-transfer radical polymerization to provide polymer layers of 2.5-, 5- and 10-nm thickness. Lipid bilayers composed of POPC (1-palmitoyl-2-oleoyl-sn-glycero-3- phosphocholine) were formed by vesicle fusion onto bare fused silica and onto each of the polyacrylamide layers. Bilayer fluidity was assessed by the diffusion of a probe, NBD-labeled phosphatidylcholine, using fluorescence recovery after photobleaching. A transmembrane protein, the human delta-opioid receptor, was inserted into each lipid bilayer, and its ability to bind a synthetic ligand, DPDPE, cyclic[2-D-penicillamine, 5-D-penicillamine]enkephalin, was detected using single-molecule fluorescence spectroscopy by labeling this ligand with a rhodamine dye. The transmembrane protein was observed to bind the ligand for all bilayers tested. The protein's electrophoretic mobility was probed by monitoring the fluorescence from the bound ligand. The 5-nm polyacrylamide thickness gave the fastest diffusion for the fluorescent lipid probe (D 1 = 2.0(±1.2) × 10 -7 and D 2 = 1.2(±0.5) × 10 -6 cm 2/s) and also the largest electrophoretic mobility for the transmembrane protein (3 × 10 -8 cm 2/V·s). The optimum in polymer thickness is suggested to be a tradeoff between decoupling from the substrate and increasing roughness of the polymer surface.

Original languageEnglish (US)
Pages (from-to)9644-9650
Number of pages7
JournalLangmuir
Volume21
Issue number21
DOIs
StatePublished - Oct 11 2005

ASJC Scopus subject areas

  • Materials Science(all)
  • Condensed Matter Physics
  • Surfaces and Interfaces
  • Spectroscopy
  • Electrochemistry

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    Smith, E. A., Coym, J. W., Cowell, S. M., Tokimoto, T., Hruby, V. J., Yamamura, H. I., & Wirth, M. J. (2005). Lipid bilayers on polyacrylamide brushes for inclusion of membrane proteins. Langmuir, 21(21), 9644-9650. https://doi.org/10.1021/la051116h