High Concentration Preparation And ATP-Responsive Assembly Of Wireframe DNA Polyhedra

Deoxyribonucleic acid（DNA）is an important carrier for storing and transmitting the genetic information of life.In addition,DNA can also be regarded as a chain polyanionic compound covalently linked by nucleotides,which has the characteristics of programmable base sequence and highly specific base pairing,and can be used as the assembly element of micro/nano materials.In the 1980s,Professor Nadrian C.Seeman of New York University put forward the idea of DNA self-assembly,hoping to help protein crystallization by constructing 3D DNA crystals in order to carry out X-ray diffraction crystallography research,which opened the field of DNA nanotechnology.After 40 years of development,DNA nanotechnology has grown into an emerging frontier field with highly interdisciplinary disciplines such as chemistry,biology,information and materials.So far,a variety of DNA self-assembly strategies have been developed,such as DNA module self-assembly,DNA origami and DNA single strand lattice assembly.Among them,DNA module self-assembly has unique advantages in two-dimensional array and three-dimensional framework polyhedron assembly due to its simplicity（only a few DNA strands）and high efficiency.In this paper,the following two studies have been carried out on the module based on DNA framework polyhedron self-assembly:1.A new method for preparing DNA framework polyhedron with high concentration and high yield was developed.Using DNA module self-assembly strategy to construct framework polyhedron is simple and efficient,but it usually needs to be assembled at low DNA concentration to reduce by-products.In order to meet the requirements of sample concentration in characterization and application research,it is often necessary to concentrate the assembled samples with low concentration,which not only increases the complexity of the experimental operation,but also causes the agglomeration loss of DNA samples.Therefore,the development of a method that can directly prepare DNA framework polyhedron with high concentration and high yield is of great significance to promote its application.In this work,by replacing the DNA self-assembly buffer solution and using monovalent cation（Na⁺）instead of divalent cation(Mg²⁺),the direct assembly of DNA framework polyhedron with high concentration and high yield was realized.It is found that:（1）framework polyhedron with concentration up toμM can be directly prepared by using Na⁺.When the concentration of DNA module is 5μM,the assembly yield is still as high as 87%,while it is 23%under Mg²⁺;（2）This strategy is suitable for module self-assembly at various viscous ends.The self-assembly of modules with different bonding strength such as 1-GC,2-GC,3-GC,4-GC and 7-nt was tried,which can achieve high-yield assembly atμM concentration;（3）This strategy is expected to be applicable to the high concentration assembly of various frame polyhedrons.Three branch module and four branch modules were used to construct DNA tetrahedron and octahedron structures with high yield atμM concentration respectively.The analysis shows that due to the polyanion property of DNA molecules,cations are needed to provide electrostatic stability in the process of self-assembly.The commonly used divalent cation Mg²⁺has strong electrostatic effect,which is not conducive to the self-correcting process of“assembly-disassembly-reassembly”between modules.Therefore,concentration related kinetic factors lead to low assembly yield at high DNA concentration.Na⁺provides appropriate electrostatic action,which can not only stabilize the DNA self-assembly structure,but also facilitate the self-error correction assembly of DNA modules.Therefore,it can effectively reduce the influence of dynamic factors and realize high-yield assembly at high concentration.This work provides a simple and efficient method for the high concentration preparation of module based on DNA framework polyhedron by replacing the cations in the self-assembly buffer,which is expected to promote the wide application of DNA polyhedron.2.The ATP responsive assembly of DNA framework polyhedron was realized based on allosteric strategy.The development of stimulus responsive dynamic self-assembly system is of great significance for the development of new nano machines and the construction of new sensors.In this work,a new DNA three branch module is designed,in which the aptamer sequence of ATP is introduced,and the structure of ATP stimulation response control module is used to realize the responsive control of DNA framework polyhedron assembly process.The results show that:（1）the initial binding of aptamer region is very important for the control of ATP stimulation response.When the initial binding effect is too strong and there is no stimulus response,that is,it can be assembled with or without ATP;When the initial binding effect is too weak,there is no stimulus response,that is,there is no assembly with or without ATP;Only proper initial binding can realize ATP stimulation responsive assembly;（2）Cations have a great influence on the selectivity of ATP stimulation response,and monovalent cation Na⁺can increase the selectivity of ATP stimulation response.In this work,a DNA framework polyhedron stimulus response assembly method is developed by using allosteric strategy,which is expected to provide a useful reference for DNA polyhedron based nano machines,drug delivery,analytical sensing and so on.