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

A novel electrochemical biosensing method for protein kinase (PKA) activity was demonstrated by using a reduced graphene oxide-polydopamine-silver nanoparticle-Ti (rGO-PDA-AgNPs-Ti) nanocomposite. The obtained nanocomposite possessed an integral capability for phosphopeptide recognition and signal readout. The polydopamine modified reduced graphene oxide (rGO-PDA) was firstly prepared based on 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 Ti cations were attached onto the PDA layer for phosphopeptide recognition according to the strong coordination ability of PDA with Ti and phosphate group. The prepared rGO-PDA-AgNPs-Ti nanocomposites were characterized with different methods. The developed rGO-PDA-AgNPs-Ti nanocomposites were then employed for electrochemical analysis of PKA-catalyzed kemptide phosphorylation. The sensitive detection toward PKA activity was realized with an experimental detection limit of about 0.01 U/mL. It may be also extended for the inhibitor evaluation. Thus, it provided a facile and sensitive means for electrochemical analysis of PKA activity and inhibitor screening.