Construction of Protein Probe with a His‐tag and an Electron‐transfer Peptide for a Target Protein Sensing

Abstract

AbstractA protein probe with an electron‐transfer peptide and a His‐tag was designed to electrochemically sense a target protein. We selected tyrosine‐rich (Y4C) and tryptophan‐rich (W4C) peptides for use as electron‐transfer agents. The peak for oxidation was based on the oxidations of the phenolic hydroxy groups in Y4C and on the indole rings in W4C. Asialofetuin (ASF) with galactose residues was the protein probe, and a galactose recognition protein, soybean agglutinin (SBA), was the target protein. A protein probe composed of an amino acid and carbohydrate residue was expected to be biocompatible. When voltammetric measurements were performed using a glassy carbon electrode, the oxidation peaks of H6Y4C and ASF‐H6Y4C appeared at the same potential. The peak current of ASF‐H6Y4C was 4‐fold that of H6Y4C because of the stronger adsorption of ASF‐H6Y4C onto the electrode. The electrode response of ASF‐H6Y4C with SBA was half that of ASF‐H6Y4C alone. By contrast, the peak current of ASF‐Y4CH6 was higher than that of ASF‐H6Y4C, which was the result of a greater degree of contact between the Y4C moieties and an electrode. On the other hand, the voltammetric behaviors of ASF with W4C and a His‐tag were similar to those with Y4C and a His‐tag. The sensitivity of SBA using ASF‐Y4CH6 was at the 10−13 M level. To confirm the function of the sensing system, measurements were performed in human serum with SBA and ASF‐Y4CH6. When SBA was added, the serum had a concentration that ranged between 5.0×10−13 and 4.0×10−12 M, and the amount of SBA that could be recovered ranged from 97 to 101%. Consequently, this system could be applied to the detection of SBA in serum.