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Addressable adsorpt...
Addressable adsorption of lipid vesicles and subsequent protein interaction studies
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- Klenkar, Goran (author)
- Linköpings universitet,Sensorvetenskap och Molekylfysik,Tekniska högskolan
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- Brian, Björn, 1980 (author)
- Linköpings universitet,Sensorvetenskap och Molekylfysik,Tekniska högskolan
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- Ederth, Thomas (author)
- Linköpings universitet,Sensorvetenskap och Molekylfysik,Tekniska högskolan
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- Stengel, Gudrun (author)
- Chalmers tekniska högskola,Chalmers University of Technology,Chalmers
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- Höök, Fredrik, 1966 (author)
- Chalmers tekniska högskola,Chalmers University of Technology,Chalmers
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- Piehler, J. (author)
- Johann Wolfgang Goethe Universität Frankfurt am Main,Goethe University Frankfurt,University of Frankfurt
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- Liedberg, Bo (author)
- Linköpings universitet,Sensorvetenskap och Molekylfysik,Tekniska högskolan
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(creator_code:org_t)
- American Vacuum Society, 2008
- 2008
- English.
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In: Biointerphases. - : American Vacuum Society. - 1559-4106 .- 1934-8630. ; 3:2, s. 29-37
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https://doi.org/10.1...
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Abstract
Subject headings
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- We demonstrate a convenient chip platform for the addressable immobilization of protein-loaded vesicles on a microarray for parallelized, high-throughput analysis of lipid-protein systems. Self-sorting of the vesicles on the microarray was achieved through DNA bar coding of the vesicles and their hybridization to complementary strands, which are preimmobilized in defined array positions on the chip. Imaging surface plasmon resonance in ellipsometric mode was used to monitor vesicle immobilization, protein tethering, protein-protein interactions, and chip regeneration. The immobilization strategy proved highly specific and stable and presents a mild method for the anchoring of vesicles to predefined areas of a surface, while unspecific adsorption to both noncomplementary regions and background areas is nonexistent or, alternatively, undetectable. Furthermore, histidine-tagged receptors have been stably and functionally immobilized via bis-nitrilotriacetic acid chelators already present in the vesicle membranes. It was discovered though that online loading of proteins to immobilized vesicles leads to cross contamination of previously loaded vesicles and that it was necessary to load the vesicles offline in order to obtain pure protein populations on the vesicles. We have used this cross-binding effect to our benefit by coimmobilizing two receptor subunits in different ratios on the vesicle surface and successfully demonstrated ternary complex formation with their ligand. This approach is suitable for mechanistic studies of complex multicomponent analyses involving membrane-bound systems.
Subject headings
- TEKNIK OCH TEKNOLOGIER -- Annan teknik (hsv//swe)
- ENGINEERING AND TECHNOLOGY -- Other Engineering and Technologies (hsv//eng)
Keyword
- HISTIDINE-TAGGED PROTEINS
- DNA
- proteins
- MATRIX
- BINDING INTERFACE
- biophysics
- molecular
- SURFACE-PLASMON RESONANCE
- lipid bilayers
- biomembranes
- ARRAYS
- adsorption
- DNA
- MICROARRAYS
- biochemistry
- surface plasmon resonance
- INTERFERON-RECEPTOR
- MEMBRANES
- I
- IMMOBILIZATION
- NATURAL SCIENCES
Publication and Content Type
- art (subject category)
- ref (subject category)
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