Construction of a sensitive electrochemical biosensor for detecting protein kinase A activity based on NH2-Zr-MOF@Thi signal amplification

Protein kinase A (PKA) plays a significant role in the regulation of biological activities by the phosphorylation of the specific proteins or substrate peptides. The highly sensitive and reusable electrochemical biosensor has been developed to detect PKA activity based on the specific coordination between Zr 4+ and phosphate group. In this strategy, the specially designed substrate peptide can be anchored on the surface of gold electrode by Au-S bond. In the presence of adenosine triphosphate (ATP) and PKA, the substrate peptide is phosphorylated. Due to the large surface area of MOF and rich Zr 4+ active sites, signal molecule Thionine (Thi) can be absorbed into the cavity of NH 2 -Zr-MOF by the electrostatic interaction to form NH 2 -Zr-MOF@Thi signal probe. The phosphorylated peptide is linked to NH 2 -Zr-MOF@Thi probe because of the strong coordination interaction between Zr 4+ and phosphate group. Therefore, the number of Thi close to the electrode surface is increased and the response signal experiences amplification, markedly enhancing the sensitivity of the detection of PKA activity, which provides an excellent electrochemical sensing platform for PKA activity detection. The response signal of the electrochemical biosensor is proportional to the logarithm of PKA concentration, ranging from 0.05 U mL −1 to 100 U mL −1 , and the limit of detection of this biosensor for PKA is 0.017 U mL −1 when the signal-to-noise ratio is 3. In particular, the proposed method can selectively distinguish the target from other interferents, and can be used to screen potential small molecule inhibitors and detect PKA activity in complex cell samples. Therefore, the developed electrochemical biosensor can be used as a new way for the determination of PKA activity and the screening of its inhibitors in the future.