Copper nanoclusters/polydopamine nanospheres based fluorescence aptasensor for protein kinase activity determination

Area of Science:

• Biochemistry
• Nanotechnology
• Analytical Chemistry

Background:

• Protein kinase activity is crucial in cellular signaling pathways.
• Accurate detection of kinase activity is essential for understanding disease mechanisms and drug development.
• Existing methods for kinase activity detection can be complex or lack sensitivity.

Purpose of the Study:

• To develop a novel fluorescence aptasensor for sensitive detection of protein kinase (PKA) activity.
• To utilize copper nanoclusters (CuNCs) and polydopamine nanospheres (PDANS) for PKA detection.
• To explore the application of the aptasensor in kinase inhibitor screening and biological sample analysis.

Main Methods:

• Construction of a fluorescence aptasensor using CuNCs and PDANS.
• Exploiting π-π stacking interactions for aptamer immobilization.
• Utilizing fluorescence resonance energy transfer (FRET) for signal transduction.
• Monitoring fluorescence changes upon PKA-mediated conversion of ATP to ADP.

Main Results:

• The aptasensor demonstrated sensitive detection of PKA activity in the range of 0.05-4.5 U/mL.
• A low detection limit of 0.021 U/mL was achieved for PKA.
• The aptasensor showed feasibility for kinase inhibitor screening using H-89 as a model.
• Successful determination of PKA activity in HepG2 cell lysates was achieved.

Conclusions:

• The developed fluorescence aptasensor provides a convenient and sensitive method for PKA activity detection.
• The aptasensor platform holds promise for high-throughput screening of kinase inhibitors.
• This approach offers a valuable tool for biochemical research and clinical diagnostics involving kinase activity.