Construction of Double-enzyme Complexes with DNA Framework Nanorulers for Improving Enzyme Cascade Catalytic Efficiency.

Efficient biocatalytic cascade reactions play a crucial role in guiding intricate, specific and selective intracellular transformation processes. However, the catalytic activity of the enzyme cascade reaction in bulk solution was greatly impacted by the spatial morphology and inter-enzyme distance. The programmability and addressability nature of framework nucleic acid (FNA) allows to be used as scaffold for immobilization and to direct the spatial arrangement of enzyme cascade molecules. Here, we used tetrahedral DNA framework (TDF) as nanorulers to assemble two enzymes for constructing a double-enzyme complex, which significantly enhance the catalytic efficiency of sarcosine oxidase (SOx)/horseradish peroxidase (HRP) cascade system. We synthesized four types of TDF nanorulers capable of programming the lateral distance between enzymes from 5.67 nm to 12.33 nm. Enzymes were chemical modified by ssDNA while preserving most catalytic activity. Polyacrylamide gel electrophoresis (PAGE), transmission electron microscopy (TEM) and atomic force microscopy (AFM) were used to verify the formation of double-enzyme complex. Four types of double-enzyme complexes with different enzyme distance were constructed, in which TDF26(SOx+HRP) exhibited the highest relative enzyme cascade catalytic activity, ~3.11-fold of free-state enzyme. Importantly, all the double-enzyme complexes demonstrate a substantial improvement in enzyme cascade catalytic activity compared to free enzymes.

Programming Intracellular Clustering of Spiky Nanoparticles via Liposome Encapsulation.

The intracellular clustering of anisotropic nanoparticles is crucial to the improvement of the localized surface plasmon resonance (LSPR) for phototherapy applications. Herein, we programmed the intracellular clustering process of spiky nanoparticles (SNPs) by encapsulating them into an anionic liposome via a frame-guided self-assembly approach. The liposome-encapsulated SNPs (lipo-SNPs) exhibited distinct and enhanced lysosome-triggered aggregation behavior while maintaining excellent monodispersity, even in acidic or protein-rich environments. We explored the enhancement of the photothermal therapy performance for SNPs as a proof of concept. The photothermal conversion efficiency of lipo-SNPs clusters significantly increased 15 times compared to that of single lipo-SNPs. Upon accumulation in lysosomes with a 2.4-fold increase in clustering, lipo-SNPs resulted in an increase in cell-killing efficiency to 45% from 12% at 24 µg/mL. These findings indicated that liposome encapsulation provides a promising approach to programing nanoparticle clustering at the target site, which facilitates advances in the development of smart nanomedicine with programmable enhancement in LSPR.