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Scientific research shows that more than 50% of proteins in the human body exist in the form of glycoproteins. As the most important post-transformation modification form of proteins in organisms, protein glycosylation has important biological functions. It not only affects the spatial structure, biological activity, transport and positioning of proteins, but also participates in molecular recognition in a variety of diseases, such as tumors (FDA has approved biomarkers for the diagnosis and monitoring of cancer diseases, most of which are glycoproteins), neurodegeneration, cardiovascular disease, suspected, immune screening and infectious diseases. The occurrence and development of diseases are accompanied by the occurrence of abnormal protein glycosylation and sugar chain structure . In addition, glycosylation also significantly affects the biological activity, biological targets, transportation, serum half-life, clearance and recognition of therapeutic agents. For example, the pharmacokinetics and effects of recombinant erythropoietin are severely affected by its glycosylation state , and the ability of monoclonal antibodies to mediate efficacy through ADCC is affected by the fucose content in the Fc region. Therefore, protein glycosylation research is another potential life research field in science after nucleic acids and proteins, with important theoretical and application significance. FDA requires that all types of glycoproteins need to be analyzed for glycoforms .

Peptide N-glycosidase F ( PNGase F, EC 3.5.1.52) is an endoglycosidase that promotes N-linked enzymes from glycoproteins. It is an important tool for glycosylation research and is widely used in the study of N- linked sugar chain sites, structural information and functions on therapeutic proteins. Rhinogen ® PNGase F (Glycerol-free) is a recombinant PNGase F from cellulose BL21. It can completely release all N-linked sugar chains (except core fucose containing α-1,3 linkages, which are common in plant and insect glycoproteins) from the protein molecules to which they are connected. It can cut between the middle N-nucleotide amide ( GlcNAc ) and asparagus starch residue ( Asn ) in the high mannose, hybrid and complex oligosaccharide parts of N-linked glycopeptides or glycoproteins to release complete oligosaccharide chains, as shown in Figure 1A. While releasing the sugar chains , the asparagine residues on a set of characteristic sequences are converted to asparagine residues, which contain a protein + 1Da mass change, as shown in Figure 1B. It is active on a variety of substrates, including blocked glycoproteins and complex mixtures of human serum glycoproteins. In addition, due to the presence of triglycerides, the enzyme is compatible with HPLC and most workflows under non-denaturing conditions. Phosphate, sulfate, and native acid groups attached to N-linked oligosaccharides do not affect the release of N-glycans .