Construction And Functional Exploration Of Complex Nano-confined Environment Based On DNA Origami

Intracellular confined environment plays an important role in maintaining the basic cellular functions such as regulating the pathways of biomolecular assembly and post-modification,providing specific conditions for biochemical reactions and directing precise transport of substances/enegy.The intracellular confined environment can be defined as the macromolecules in the cytoplasm confining biological reactions,membranes confining membrane proteins or organelles confining series related metabolic processes to realize certain functions.Two strategies to simulate the intracellular confined environment in vitro have been proposed: macro-scale construction and micro/nano-scale construction.A variety of mimicking systems in micro/nano scale have been well developed during last decades,such as vesicles,micelles and microgels.However,it is still a major challenge to accurately control the size of the confined environment and precisely regulate the number and distribution of the molecules in it.Based on the development of structural DNA nanotechnology,DNA origami has been widely used to build a variety of nano-scale structures.In this thesis,we have applied DNA origami to construct complex confined environment with precise size,and further explored the behavior of both enzymatic reaction and molecular assembly in confined environment.Firstly,a switchable DNA origami nanochannel has been constructed with a responsive nano-confined environment.The DNA nanochannel was designed with a shutter at one end,which could be reversibly switched between open and close state,through the DNA chain exchange reactions.The opening and closing of the shutter influenced the transport of materials through this nanochannel and the processes have been visualized by a GOx-HRP enzymatic cascade reaction.To further reveal the mechanism,taking advantage of the designability of DNA origami,confined nanochannels with different channel size have also been designed and investigated.Secondly,we have constructed a nanomembrane-confined environment through the frame-guided assembly on the basis of 2D DNA origami template.The nanomembrane is constituted by the dendrimer-based molecule(DDOEG).Lipases are embedded in the membrane,whose number and position are precisely controlled by the design of DNA origami.This confined nanomebrane could enrich the hydrophobic substrate to the enzymes,which has improved the efficiency,comparing with the free enzymes.Due to the composition of the membrane,substrate selectivity has been observed: only those hydrophobic molecules with aromatic groups could be enriched.It was also been proved that this confined membrane could protect the embedded enzymes from protease degradation.At last,we have constructed hydrophobic confined template through DNA origami and DDOEG to direct the assembly of DNA-poly(propylene oxide)within it.The assembly exhibits a micellar structure in the confined environment,with a size that matches the nanochannel port.Through tuning the design of DNA origami,we can accurately adjust the position of assembly in the nanochannel.Furthermore,it has been observed that the assembly in the confined nanochannel could also enrich orthosilicate in solution and further controls the growth of silica in situ.