Solution Paramagnetic Relaxation Enhancement To Study The Structure Of RNA And Its Dynamic Changes

RNA is a single-stranded molecule formed by transcription of DNA as a template.It is both an information molecule and a functional molecule.In addition to transmitting information as a genetic material,RNA also plays an important catalytic and regulatory role in life activities such as gene expression and signal transduction.The diversity of structures determines the diversity of functions.However,based on the number of structures of proteins in PDB,the structure of RNA is less than 1%,which is directly related to the lack of research methods caused by RNA structural flexibility.On the other hand,with the development of biology,structural research is no longer limited to static structures.For flexible single-stranded RNA,dynamic structure research is also extremely important.Therefore,it is of great significance to study the structure of RNA and its dynamic changes.Common methods for studying RNA structures include X-ray crystal diffraction,nuclear magnetic resonance,and cryo-electron microscopy.Due to the fact that RNA is relatively dynamic and difficult to crystallize and has a relatively small molecular weight,high-resolution structures cannot be obtained by X-ray crystal diffraction or cryo-electron microscopy,but the nuclear magnetic resonance method can not only perform experiments in a solution under physiological conditions,but also structural changes were observed by measuring relevant parameters during dynamic changes.Solution paramagnetic relaxation enhancement(sPRE)is a nuclear magnetic resonance method that can obtain long-range distance information in the 40 ? range.Compared with the conventional PRE,sPRE distributes the paramagnetic probe evenly in the solution,avoiding structural changes caused by the probe label,and is especially suitable for RNA molecules that are difficult to achieve labeling.Therefore,this paper intends to use the sPRE method to study the structure of RNA and its dynamic changes.In this paper,the preparation method of in vitro transcription and solid phase synthesis was used to obtain high-purity RNA samples that meet the requirements.Then the sPRE method was used to analyze the structure of the model molecule UUCG tetraloop,and the feasibility of sPRE in RNA structure analysis was verified.Further applied to the structural study of MMTV A27 G with more complex pseudo-knot structure,it was found that even In the case of pairing the sPRE data of the hydrogen atom on the imino group,the structure is also accurately characterized.Finally,this method is extended to the field of RNA dynamic change research,and the relatively dynamic structural ensemble of HRV-14 cre is obtained.In summary,this paper develops the application of sPRE in RNA structure and dynamic changes,and provides new methods and ideas for RNA structure research.