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Abstract:
Reconstitution of transmembrane proteins into supported lipid bilayer (SLB) membranes has often been hampered by strong interactions of protein domains with the underlying solid support leading to loss of both activity and mobility within the plane of the lipid bilayer. Polymer cushioned SLBs can overcome this by avoiding direct contact with the support. To extend this approach we developed an anionic polymer cushion system that allows tunable lipid mobility as well as functional integration of the transmembrane protein β-amyloid precursor protein cleaving enzyme (BACE) into SLBs. Fluorescence recovery after photobleaching analysis revealed a homogeneous distribution and high lateral mobility of the reconstituted BACE in cushioned SLBs while an impaired mobility and inhomogeneous clustering of reconstituted BACE were found in SLBs on silicon oxide substrates. The cushioning of SLBs led to increased incorporation and enhanced enzymatic activity of the reconstituted BACE with a direct correlation between lipid mobility and BACE activity. The utilized polymer cushion system allows the successful reconstitution of transmembrane proteins within SLBs with tunable properties.
