Poly(gamma-methyl L-glutamate)s with Asp, Tyr, Trp, and Ser residues at the amino terminal as the binding sites in the nicotinic acetylcoline receptor and gamma-aminobutyric acid receptor, respectively, were prepared. The number average degree of polymerization of the polypeptides was 22. Dipalmitoylphosphatidylcholine monolayers containing the polypeptides were formed on buffered subphase solutions with and without substrates and were transferred onto gold-deposited glass plates. The polypeptides rearranged to form an assembly, owing to the interaction between the terminal amino acids and the substrates in the lipid monolayer on the buffer solution containing substrates. The polypeptide assembly vertically oriented to the monolayer surface. The binding ability of the substrates to the polypeptide assembly in the lipid monolayer was characterized by surface plasmon resonance measurements. The polypeptide assembly in the lipid monolayer conditioned on the acetylcholine aqueous solution exhibited a specific binding ability for the acetylcholine molecule. However, the polypeptide assembly lipid monolayer system prepared on the gamma-aminobutyric acid aqueous solution showed a specific binding ability for the gamma-aminobutyric acid. This behavior may have arisen from a substrates-induced rearrangement of the polypeptide assembly in the lipid monolayer, forming corresponding binding sites similar to that found in the nicotinic acetylcholine receptor and gamma-aminobutyric acid receptor, respectively.