Sensitive detection of dipeptidyl peptidase based on DNA–peptide conjugates and double signal amplification of CHA and DNAzymes

Abstract

Dipeptidyl peptidase IV (DPPIV) is an enzyme belonging to the type II transmembrane serine protease family that has gained wide interest in the fields of hematology, immunology, and cancer biology. Moreover, DPPIV has emerged as a promising target for therapeutic intervention in type II diabetes. Due to its biological limitations, traditional strategies cannot meet the requirements of low abundance DPPIV analysis in complex environments. In this work, combining the high programmability of DNA and the chemical diversity of peptides, we designed DNA–peptide conjugates that can be specifically recognized, polypeptides as specific substrates for target DPPIV and DNA probes as primers for catalytic hairpin assembly (CHA), recycling a large amount of DNAzymes by triggering CHA amplification. The DNAzyme substrate modified with the FAM fluorescent group was immobilized on the surface of gold nanoparticles by S–Au chemical bonds to form a signal output probe. The DNAzymes enzyme cleaved the substrate of the signal outputs probe, yielding a double-amplified fluorescence signal. This method has a detection limit as low as 0.18 mU mL⁻¹ and a linear range of 0–5 mU mL⁻¹ in serum samples, showing high stability and good potential for practical applications.