Single-Molecule Imaging of Enzymatic Reactions on DNA Origami

The interaction between enzymes is very important for understanding of enzyme functions and shed light on enzymatic reaction mechanisms. With the development of DNA nanotechnology, DNA origami has become a powerful tool for analyzing the dynamic behavior of enzyme molecules. In this protocol, a method for imaging and analysis of single-molecule cascade enzyme reactions on DNA origami raft by total internal reflection fluorescence microscopy (TIRFM) is described. Through trajectory analysis and calculation, the diffusion of downstream enzymes in enzymatic reaction and chemotaxis of enzymatic reactions were elucidated at the single molecular level.