Potential-assisted enzymatic peptide synthesis in aqueous solution has been performed by cx-chymotrypsin immobilized in conducting polymer polypyrrole matrix. Chymotrypsin is electrochemically adsorbed on a Pt electrode, and then a thin polypyrrole membrane is electrochemically prepared on the enzyme layer. The enzyme/polypyrrole on an electrode retains chymotrypsin activity and the equilibrium of the enzymatic reaction shifts to the synthesis by ca. 25% due to the hydrophobic environment caused by polypyrrole matrix. Here, Ac-Phe-OEt and Ala-NH2 are used as model substrates of acyl donor ester and a nucleophile acceptor, respectively, to synthesize Ac-Phe-AlaNH(2). Both the synthetic and the hydrolytic activity of the immobilized chymotrypsin are found to be enhanced several times when a positive potential (0.4-0.8 V vs Ag/AgCl) is applied to the electrode. In addition, the equilibrium of the enzymatic reaction shifts further to synthesis. About 73% of synthetic yield of Ac-Phe-Ala-NH2 is obtained from equimolar concentration (200 mM) of Ac-Phe-OEt and Ala-NH2 in aqueous solution. The effects of solution pH, temperature, organic solvent concentration, and substrates concentration on the peptide synthesis are also described. An explanation of possible effects of the applied voltage which causes the remarkable increase of catalytic activity is also presented.