Affinity Capturing and Surface Enrichment of a Membrane Protein Embedded in a Continuous Supported Lipid Bilayer
Journal article, 2016

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.

surface chemistry

membrane proteins

His-tag

supported lipid bilayer

affinity purification

Author

Anders Gunnarsson

AstraZeneca AB

Lisa Simonsson Nyström

Chalmers, Physics, Biological Physics

Sabina Burazerovic

Chalmers, Physics, Biological Physics

J. Gunnarsson

AstraZeneca AB

Arjan Snijder

AstraZeneca AB

S. Geschwindner

AstraZeneca AB

Fredrik Höök

Chalmers, Physics, Biological Physics

ChemistryOpen

2191-1363 (eISSN)

Vol. 5 5 445-449

Subject Categories

Biochemistry and Molecular Biology

DOI

10.1002/open.201600070

PubMed

27777836

More information

Latest update

3/21/2018