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Detecting Spontaneously Inserting Transmembrane Peptides and Proteins in Tethered Phospholipid Bilayers

Published

Author(s)

John T. Elliott

Abstract

We have fabricated a tethered phospholipid bilayer sensor platform for detecting the function of spontaneously inserting transmembrane peptides and proteins. A mixed [beta]-mercaptoethanol/1-thia-hexa(ethyleneoxy)-octadecanoate monolayer was initially assembled on a planar gold surface. The ethyleneoxide portion of the amphipathic anchor was designed to provide a thin aqueous layer between the solid support and the membrane for accommodating small extramembranous regions of transmembrane proteins. Impedance analysis of a 1-palmitoyl-2-oleoyl-phosphotidylcholine (POPC) membrane formed over the mixed monolayer by ethanol painting methods was consistent with the formation of a bilayer (Cm[approximately equal to]0.7 [mu]F/cm2) structure over a hydrated aqueous layer ([approximately equal to] 9 [mu]F/cm2). The bilayer was highly insulating and defect free as determined by AC impedance analysis in the presence of soluble FeCN63/4- redox couple. Addition of the peptide toxin, melittin, resulted in the formation of pores large enough to allow FeCN63/4- ions access to the gold electrode. We are now using simultaneous surface plasmon resonance and impedance measurements to further characterize the physical structure of the tethered membrane and the pores that are formed by melittin. We are also extending these studies to detect the presence of the spontaneously inserting pore forming protein, [alpha]-hemolysin ([alpha]H). Reconstitution of [alpha]H into a biosensor platform may allow us to detect DNA oligomers that alter the conducting properties of the [alpha]H pore.
Citation
Detecting Spontaneously Inserting Transmembrane Peptides and Proteins in Tethered Phospholipid Bilayers

Keywords

DNA oligomers, impedance measurements, melittin, surface plasmon resonance, tethered phospholipid bilayers, transmembrane peptides, transmembrane proteins

Citation

Elliott, J. (2001), Detecting Spontaneously Inserting Transmembrane Peptides and Proteins in Tethered Phospholipid Bilayers, Detecting Spontaneously Inserting Transmembrane Peptides and Proteins in Tethered Phospholipid Bilayers (Accessed April 16, 2024)
Created February 1, 2001, Updated February 17, 2017