A new species was formed when protein P-23 (one segment of ciliate Euplotes octocarinatus centrin) was added to a solution of Eu3+. The interaction between P-23 and Eu3+ was investigated by cyclic voltammetry, pulse voltammetry and electrochemical impedance spectroscopy in 10 mM N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid (HEPES) buffer (pH 7.4) using a pyrolytic graphite electrode. The formal potential (E degrees') of Eu3+ shifted from -0.61 to -0.84 V (versus saturated calomel electrode) after P-23 was added to the Eu3+ solution. The diffusion coefficient (D), the charge-transfer coefficient (alpha) and the electron transfer standard rate constant (k(s)) were obtained in the absence and the presence of P-23. The affinity constant of Eu3+ and P-23 was determined to be (1.89 +/- 0.51) x 10(4) M-1. The electrochemical investigation of europium bound to the protein provided useful data for the studies of calcium-binding proteins. (C) 2010 Elsevier Ltd. All rights reserved.