Porous alumina substrates with a pore diameter of 60 nm were functionalized with cholesterylpolyethylenoxy thiol (CPEO3) chemisorbed on gold, which is a prerequisite for the formation of pore-suspending membranes obtained from spreading of large unilamellar vesicles with a mean diameter of 730 nm. By means of vesicle fusion, a large number of gramicidin D ion channels, selective for monovalent cations, was successfully reconstituted into these pore-suspending membranes as probed by impedance spectroscopy. An electrical equivalent circuit was developed to fully describe the electrical properties of the gramicidin D-doped membrane system, which allowed us not only to extract the membrane resistance but also the mass transport in bulk solution as a function of the monovalent cation concentration in solution. Blocking of channel activity was demonstrated by the addition of Ca2+ ions verifying the reconstitution of fully functional peptides.
