An Integral Recognition and Signaling for Electrochemical Assay of Protein Kinase Activity and Inhibitor by Reduced Graphene Oxide-Polydopamine-Silver Nanoparticle-Ti4 Nanocomposite

摘要

A novel electrochemical biosensing method for protein kinase (PKA) activity was demonstrated by using a reduced graphene oxide-polydopamine-silver nanoparticles-Ti4 (rGO-PDA-AgNPs-Ti4 ) nanocomposite, which interestingly served as an integral phosphopeptide-recognizing and signal reporting platform. The polydopamine modified reduced graphene oxide (rGO-PDA) was first prepared by a self-polymerization method of dopamine. The silver ions were adsorbed onto polydopamine (PDA) layer and directly reduced into silver nanoparticles (AgNPs), which was used for electrochemical signal reporting. Then, the Ti4 cations were attached onto the PDA layer for phosphopetide recognition according to the strong coordination ability of PDA with Ti4 and phosphate group. The prepared rGO-PDA-AgNPs-Ti4 nanocomposites were well-characterized by various techniques. With the full advantage of specific recognition of Ti4 for the phosphate group, electrochemical response of AgNPs, and excellent conductivity of rGO, the rGO-PDA-AgNPs-Ti4 nanocomposite endowed a label-free and one-step electrochemical analysis of kemptide phosphorylation catalyzed by PKA. The detection limit for PKA activity was experimentally achieved as 0.01 U/mL, which was evidently lower than most of the reported methods. The proposed sensing strategy could be also applied for an efficient inhibitor evaluation. Therefore, it offered an excellent pathway for a generic and sensitive electrochemical assay of PKA activity and inhibitor.