AVS 50th International Symposium
    Biomaterial Interfaces Wednesday Sessions
       Session BI-WeP

Paper BI-WeP9
Multilayers of Functionalized Liposomes for Improved SPR Analysis of Transmembrane Proteins

Wednesday, November 5, 2003, 11:00 am, Room Hall A-C

Session: Poster Session
Presenter: A. Granéli, Chalmers University of Technology, Sweden
Authors: A. Granéli, Chalmers University of Technology, Sweden
F. Höök, Chalmers University of Technology, Sweden
Correspondent: Click to Email
The cell membrane consists of a large fraction of transmembrane proteins, which mediates and performs a large number of reactions taking place in the cell membrane or at the cell membrane surface. All transmembrane proteins consist of a hydrophobic part that transverse the bilayer, which make most of them insoluble in water and therefore difficult to study. Accordingly, functional studies of individual transmembrane proteins generally require dissolving or reconstituting procedures, such as the use of detergents or incorporation in lipid assemblies such as proteoliposomes. In biosensing applications, including drug screening and medical diagnostics, as well as for fundamental studies of transmembrane proteins, surface-based techniques have turned out to be important analytical tools. Application of such techniques require that the proteins are immobilized on a solid surface, which often tends to have a negative influence on the protein activity. The necessity of having the transmembrane proteins residing in lipid membranes complicates immobilization of sufficient amounts of protein. To allow the use of surface analytical tools for studies of transmembrane proteins, protocols that enhance the amount of immobilized protein, thus the signal, are required. For that purpose, we have developed a strategy where multilayers of proteoliposomes are immobilized on Au or SiO@sub 2@ surfaces, proven versatile for studies of ligand-interaction kinetics using the quartz crystal microbalance with dissipation monitoring (QCM-D) and surface plasmon resonance (SPR) techniques. This was achieved by utilizing a DNA-modified surface, to which proteoliposomes modified with complementary DNA was immobilized; a process that was possible to repeat up to at least 6 layers, thus allowing the use of the full sensing depth of QCM-D and SPR. Signal-amplification using the liposome multilayer approach was proven via dissociation or binding from/to the transmembrane protein transhydrogenase.