DNA flower-encapsulated horseradish peroxidase with enhanced biocatalytic activity synthesized by an isothermal one-pot method based on rolling circle amplification.

DNA nanotechnology has been developed to construct a variety of functional two- and three-dimensional structures for versatile applications. Rolling circle amplification (RCA) has become prominent in the assembly of DNA-inorganic composites with hierarchical structures and attractive properties. Here, we demonstrate a one-pot method to directly encapsulate horseradish peroxidase (HRP) in DNA flowers (DFs) during RCA. The growing DNA strands and Mg2PPi crystals lead to the construction of porous DFs, which provide sufficient interaction sites for spontaneously incorporating HRP molecules into DFs with high loading capacity and good stability. Furthermore, in comparison with free HRP, the DNA flower-encapsulated HRP (termed HRP-DFs) demonstrate enhanced enzymatic activity, which can efficiently biocatalyze the H2O2-mediated etching of gold nanorods (AuNRs) to generate distinct color changes since the longitudinal localized surface plasmon resonance (LSPR) frequency of AuNRs is highly sensitive to the changes in the AuNR aspect ratio. Through rationally incorporating the complementary thrombin aptamer sequence into the circular template, the synthesized HRP-DF composites are readily used as amplified labels for visual and colorimetric detection of thrombin with ultrahigh sensitivity and excellent selectivity. Therefore, our proposed strategy for direct encapsulation of enzyme molecules into DNA structures shows considerable potential applications in biosensing, biocatalysis, and point-of-care diagnostics.