Abstract
While protein kinases play a pivotal role in various fundamental biological processes, their overexpression can lead to several human diseases. Herein, we propose a highly sensitive leaky surface acoustic wave (LSAW) biosensor for detecting protein kinase A (PKA) activity based on peptide–templated copper nanoclusters for signal enhancement. The substrate peptide P1, specific for PKA, was self–assembled on a gold-coated SAW surface. In the presence of PKA and adenosine-5′-triphosphate (ATP), the serine within P1 was phosphorylated. Another peptide, P2, which contained a cluster formation sequence CCY, was used to prepare the peptide-templated copper nanoclusters (P2-CuNCs) and a guanidine group within arginine residue was used to identify the phosphorylation site specifically. The P2-CuNCs were captured on phosphorylated peptide P1 through covalent-like the interaction of guanidine–phosphate, which resulted in amplified SAW signals. Under the optimal conditions, the SAW sensor showed a quantitative readout to PKA concentration in the range 0−50 U/mL with a detection limit of 0.02 U/mL for PKA. In addition, the biosensor was used to screen protein kinase inhibitors and detect protein kinase activity in drug–stimulated complex cell lysates, showing great potential use in drug discovery and disease diagnosis.
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•We reported a novel leaky surface acoustic wave biosensor for detection of PKA activity.•Versatile peptide–templated copper nanoclusters (P2–CuNCs) were employed for highly sensitive detection of PKA.•The SAW sensor showed high sensitivity, with a detection limit of 0.02 U/mL for PKA.•The biosensor was used to detect protein kinase activity in drug-stimulated complex cell lysates.