Construction And Application Of Nano-fluorescent Probes For Simultaneous Recognition Of Protein Phosphorylation And Glycosylation Sites

Post-translational modification of proteins involves almost all life processes and plays an important regulatory role in the function and activity of proteins.Among them,protein glycosylation and phosphorylation are the most common post-translational modifications.As a biomarker and therapeutic target,glycosylation sites are involved in the occurrence and development of many diseases.Recent studies have shown that abnormal levels of phosphoprotein are closely related to the pathogenesis of many diseases.However,it is still not clear whether there is a certain correlation between glycosylation and phosphorylation of proteins in biology.Therefore,studying the relationship between protein glycosylation and phosphorylation is considered to be an effective way to understand the occurrence and development of major diseases,which is of great significance for studying the pathogenesis and early diagnosis of diseases.Based on this,two types of MOFs nanoprobe have been developed respectively with Zr(IV)and boric acid as active centers.Fluorescence detection of phosphorylation sites was achieved through the specific interaction of Zr(IV)with phosphate.Utilizing the specific effect of boric acid and cis diol,the alizarin red was used to regulate the fluorescence of the probe,and the specific recognition of the glycosylation site was achieved.Finally,the prepared nanoprobe was used for in situ fluorescence imaging of glycosylation and phosphorylation levels in depressed mice.The specific work is as follows:1.Using Zr(IV)and boric acid as active centers,we designed and synthesized MOFs nanoprobes that recognize both glycosylation and phosphorylation sites.Using the specific interaction between Zr(IV)and phosphate,fluorescence imaging of phosphorylation sites was achieved.In order to achieve specific recognition of glycosylation sites,the boronic acid group was modified in the MOFs structure,and fluorescence imaging of glycosylation sites was achieved by regulating the competition between alizarin red and glycosylation sites.Subsequently,a mouse model of depression was established and experimentally verified by a behavioral test.Finally,the relationship between glycosylation and phosphorylation in depressive mice was studied by a combination of small animal in vivo imaging and two-photon imaging.The results showed that the levels of glycosylation and phosphorylation sites in depression model mice were significantly lower than those in normal mice.This study provides a good fluorescent tool for further revealing the relationship between glycosylation and phosphorylation and its mechanism of disease.2.Based on previous experiments,5,10,15,20-tetrakis(4-carboxyphenyl)porphyrin with strong two-photon properties was used to regulate the fluorescence of the probe.MOFs-based nanoprobes were prepared and used for glycosyl groups.Simultaneous recognition of phosphorylation and phosphorylation sites.The coordination between the phosphate and the zirconium metal node interrupts the charge transfer between the ligand molecule and the zirconium metal node,so that the fluorescence of 2-aminoterephthalic acid and porphyrin is restored,and the paraphosphoric acid is realized.The detection of chemical sites due to the strong two-photon nature of porphyrins can effectively avoid the interference of background autofluorescence in cell experiments and animal experiments.At the same time,the boronic acid group was modified in the structure of MOFs to realize the fluorescence detection of glycosylation sites.Finally,the probe was applied to the levels of phosphorylation and glycosylation in depression mice before and after fluoxetine administration.This study further clarifies the relationship between protein glycosylation and phosphorylation in depression models,and provides new ideas for the study of depression mechanisms.