Zr-MOF based electrochemical biosensor coupled with [Ru(NH₃)₆]³⁺ mediated signal amplification for detection of...

Protein kinase A (PKA) is a key cellular signaling enzyme that transfers phosphate groups from adenosine triphosphate (ATP) to serine or threonine residues to regulate activity and downstream processes. This work presents a highly sensitive and selective electrochemical biosensor for PKA activity detection. Peptide and DNA are self-assembled on Au nanoparticles (Au NPs) via AuS bonds to form DNA-Au NPs-peptide complexes. The peptide is phosphorylated in the presence of PKA. Moreover, zirconium-metal organic framework (Zr-MOF) acts as an electrode substrate to facilitate charge transfer and molecular capture. Critically, phosphorylation allows complex immobilization on the electrode through ZrOP bonds between phosphorylated peptide and Zr-MOF. [Ru(NH₃)₆]³⁺ is electrostatically bound to DNA as an electroactive species for signal amplification. PKA activity regulates the amount of immobilized DNA, which determines the loading amount of [Ru(NH₃)₆]³⁺, and quantitative analysis is achieved by monitoring signal changes. The response signal of the biosensor exhibits a linear relationship with the logarithm of PKA concentration in the range of 0.05 to 100 U mL^-1, and the limit of detection is 0.016 U mL^-1. This biosensor can discriminate PKA from interferents, and allow inhibitor screening and cellular sample detection, providing a promising approach for PKA determination and inhibitor research.