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Affinity Capturing and Surface Enrichment of a Membrane Protein Embedded in a Continuous Supported Lipid Bilayer

A. Gunnarsson ; Lisa Simonsson Nyström (Institutionen för fysik, Biologisk fysik (Chalmers)) ; Sabina Burazerovic (Institutionen för fysik, Biologisk fysik (Chalmers)) ; J. Gunnarsson ; A. Snijder ; S. Geschwindner ; Fredrik Höök (Institutionen för fysik, Biologisk fysik (Chalmers))
ChemistryOpen (2191-1363). Vol. 5 (2016), 5, p. 445-449.
[Artikel, refereegranskad vetenskaplig]

Investigations of ligand-binding kinetics to membrane proteins are hampered by their poor stability and low expression levels, which often translates into sensitivity-related limitations impaired by low signal-to-noise ratios. Inspired by affinity capturing of water-soluble proteins, which utilizes water as the mobile phase, we demonstrate affinity capturing and local enrichment of membrane proteins by using a fluid lipid bilayer as the mobile phase. Specific membrane-protein capturing and enrichment in a microfluidic channel was accomplished by immobilizing a synthesized trivalent nitrilotriacetic acid (tris-NTA)-biotin conjugate. A polymer-supported lipid bilayer containing His(6)-tagged b-secretase (BACE) was subsequently laterally moved over the capture region by using a hydrodynamic flow. Specific enrichment of His(6)-BACE in the Ni2+-NTA-modified region of the substrate resulted in a stationary three-fold increase in surface coverage, and an accompanied increase in ligand-binding response.

Nyckelord: affinity purification, His-tag, membrane proteins, supported lipid bilayer, surface chemistry



Denna post skapades 2016-12-02. Senast ändrad 2017-06-28.
CPL Pubid: 245795

 

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Institutioner (Chalmers)

Institutionen för fysik, Biologisk fysik (Chalmers)

Ämnesområden

Biokemi och molekylärbiologi

Chalmers infrastruktur