Study On The Fluorescence Probes For The Tolemeric Multimeric G-quadruplex And Potato Virus Y G-quadruplex

Guanine-rich sequences may form a special nucleic acid secondary structure called G-quadruplex,which is distributed in the genome of animals,plants and microorganisms.G-quadruplexes participate in many biological processes such as replication,transcription,telomere maintenance,epigenetic regulation and other biological processes.In view of the special structure and biological function of the G-quadruplex,the design and development of fluorescent probes that can target the G-quadruplex is helpful to study the structure and biological function of the G-quadruplex.In this study,based on the structure of DNA G-quadruplex(human telomere G-quadruplex TTA45)and RNA G-quadruplex(Plant Potato Virus Y G-quadruplex),we employed bioinformatics technology and virtual screening to discover fluorescence probes.The results were listed as follows:1.The fluorescent probe that specifically recognizes the human telomeirc multimeric G-quadruplex TTA45.One of the reasons that tumor cells have the ability to proliferate indefinitely is that their telomere terminal DNA is abnormally extended under the action of reverse transcriptase.Telomere G-quadruplexes can block the reverse transcription process of telomerase to a certain extent,and telomere DNA is programmed to shorten,thus normal apoptosis of tumor cells occurs.First,based on the structure of the telomere multimeric G-quadruplex TTA45,we obtained a series of quinoline derivatives via the virtual screening.Then,we found that the BEPQ-1 exhibited specific fluorescence response to TTA45 by the fluorescence assay.Job-plot analysis verified the interaction between BEPQ-1 and TTA45 at a stoichiometric ratio of 1:1.Finally,2-Ap experiment showed that BEPQ-1 simultaneously bound to on the plane of two consecutive G-quadruplexes.By extending the length of the loop region of TTA45,we further demonstrated the binding sites.The fluorescence intensity of BEPQ-1 decreased with the longer of the loop region.UV titration experiments and glycerol experiments verified that BEPQ-1 interacted with the TTA45 G-quadruplex through ?-? stacking.2.Specific recognition of Potato Virus Y(PVY)G-quadruplex by a fluorescence probePotato Y virus,a positive-sense single-stranded RNA(+ss RNA)virus,could affect the quality and yield of many important Solanaceae plants including tomato,potato and tobacco.The design of fluorescent probes that specifically target the PVY PQS(G-quadruplex forming sequence)is conducive to the study of the structure and function of the PQS in PVY.The probe can serve as a fluorescent dye for labeling the virus,which is benefited to reveal the distribution and proliferation of potato Y virus in vivo and the design of antiviral drugs by targeting PQS.In this study,we analyzed the presence and conservation of PQSs in the PVY genome.The circular dichroism spectrum experiment proved that the sequence PQS-10 can form a G-quadruplex structure.Then,we found that the thiazole orange derivative MBA-1(TO1)showed the fluorescence selectivity for PQS-10 via fluorescence experiments.Confocal imaging results showed that TO1 colocalized with PQS-10 G-quadruplexes in live cells.We also designed and synthesized a series of derivatives based on the lock-and-key structure,It is expected that we could obtain the fluorescence probes with high specificity and selectivity for PQS-10.The structure-activity relationship based on these derivatives will contribute to the design and discovery of small molecules to specifically target RNA G-quadruplexes.