Scientists develop new tools for protein "reading sugar"

Scientists in the United States have developed a molecular tool called exoo to decode the attachment sites of specific sugars on proteins, which may be caused by diseases The research results published in molecular system biology introduced the development process of the tool and its successful application in human blood, tumor and immune cells
Half of all proteins produced by human cells have attached sugar molecules, of which N-glycan and O-glycan are the most common At present, O-glycans are difficult to be studied due to lack of tools for recognition In addition, although proteins are encoded and manufactured according to DNA blueprints, whether or not sugar molecules are attached to proteins and the number of sugar molecules attached are different, especially in the case of disease Weiming Yang, assistant researcher of pathology at Johns Hopkins University School of medicine, said: "in the field of glycobiology, the biggest challenge for researchers is to determine which sugars are bound to which proteins and where We have now developed a reliable way to determine this In addition, we have demonstrated that exoo can be used for all types of samples, including tissues, fluids and cells " The team has developed a program called operator to study other glycoconjugates and bacterial enzymes It can cleave proteins at the attachment site of O-glycans Today, they use this program to combine with a large number of different reactions to develop exoo In short, the method first digests the protein sample into smaller fragments, then connects these fragments to the solid support, after treatment with oprator enzyme, these fragments release small pieces of protein at the O-glycan attachment site, and then analyzes small pieces of protein to determine the location of sugar attachment
"It's a very good approach," says Yang We prove that exoo is the first effective tool to recognize the glycan attachment site and the specific glycan attached to the site " To determine the effectiveness of the new procedure, the team first used exoo for a well-studied glycol fetal calf protein known to contain six potential O-glycan attachment sites After testing the protein with exoo, the team identified all six known sites and found a seventh The team then looked at whether exoo could be used in larger, complex protein mixtures They used the exoo method to study normal kidney samples, cancerous kidney samples from three patients with renal clear cell carcinoma, and T cells and serum The researchers mapped 2804 O-glycan protein fragments in kidney tissue, including 1781 attachment sites from 592 proteins In T cells, there are 1982 protein fragments containing O-glycan protein, including 1295 attachment sites from 590 proteins In serum, there are 1060 protein fragments containing O-glycan protein, including 732 attachment sites from 306 proteins The researchers compared these data with data from three different glycoproteome databases completed by other research teams, and the results showed that there were 2580 unreported O-glycan sites, an increase of 94% over the known sites When the researchers compared normal and cancerous kidney tissues, they found 56 proteins of O-glycans attached to the two tissues At the same time, they found new changes in two proteins known to be unrelated to renal cancer in tumor cells compared to normal cells These results suggest that O-glycan attachment to proteins is dynamic and may be highly disease-specific, the researchers said Hu Zhang, Professor of pathology at Johns Hopkins University School of medicine, senior author of the report, said: "we hope that this tool can help researchers and scientists who study O-linked glycosylation in normal biology and diseases.