The sequential layer-by-layer formation of peptide-supported bimolecular lipid membranes at solid supports is described. In the first step, thiol-derivatized peptide sequences of 5 and 7 amino acids are assembled on a Au substrate. After activation of their COOH-terminus phospholipid molecules (DMPE) are covalently attached via an amid bond to form a tethered monolayer on the Au electrode. The different preparation steps are analyzed by Fourier transform IR, X-ray reflectometry, and surface plasmon spectroscopy. The latter technique is then also used to on-line monitor at the solid/solution interface the formation of a bilayer by fusion of vesicles prepared from a fluid Lipid mixture with and without reconstituted proteins. The obtained thicknesses and capacitance values are compatible with the tethered bilayer model and point to an incorporation of ATPase into these membrane matrices.