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Title: Droplet interface bilayers for the study of membrane proteins
Author: Hwang, William
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2008
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Aqueous droplets submerged in an oil-lipid mixture become enclosed by a lipid monolayer. The droplets can be connected to form robust networks of droplet interface bilayers (DIBs) with functions such as a biobattery and a light sensor. The discovery and characterization of an engineered nanopore with diode-like properties is enabling the construction of DIB networks capable of biochemical computing. Moreover, DIB networks might be used as model systems for the study of membrane-based biological phenomena. We develop and experimentally validate an electrical modeling approach for DIB networks. Electrical circuit simulations will be important in guiding the development of increasingly complex DIB networks. In cell membranes, the lipid compositions of the inner and outer leaflets differ. Therefore, a robust model system that enables single-channel electrical recording with asymmetric bilayers would be very useful. Towards this end, we incorporate lipid vesicles of different compositions into aqueous droplets and immerse them in an oil bath to form asymmetric DIBs (a-DIBs). Both α-helical and β-barrel membrane proteins insert readily into a-DIBs, and their activity can be measured by single-channel electrical recording. We show that the gating behavior of outer membrane protein G (OmpG) from Escherichia coli differs depending on the side of insertion in an asymmetric DIB with a positively charged leaflet opposing a negatively charged leaflet. The a-DIB system provides a general platform for studying the effects of bilayer leaflet composition on the behavior of ion channels and pores. Even with the small volumes (~100 nL) that can be used to form DIBs, the separation between two adjacent bilayers in a DIB network is typically still hundreds of microns. In contrast, dual-membrane spanning proteins require the bilayer separation to be much smaller; for example, the bilayer separation for gap junctions must be less than 5 nm. We designed a double bilayer system that consists of two monolayer-coated aqueous spheres brought into contact with each side of a water film submerged in an oil-lipid solution. The spheres could be brought close enough together such that they physically deflected without rupturing the double bilayer. Future work on quantifying the bilayer separation and studying dual-membrane spanning proteins with the double bilayer platform is planned.
Supervisor: Bayley, Hagan Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Biochemistry ; Chemistry & allied sciences ; Biophysics ; Chemical biology ; High-Throughput Screening ; Nanomaterials ; Electrical engineering ; Mathematical modeling (engineering) ; droplet interface bilayer ; protein pore ; ion channel ; protocell networks ; electrical modeling ; asymmetric bilayers ; double bilayers ; molecular computing