Protein peptides are considered to be promising biomolecules in drug development because of their low toxicity, high specificity, and good pharmacological activity against certain diseases, but their low stability, susceptibility to degradation by various enzymes in the body, and rapid renal clearance lead to short half-lives, which limit their use. Based on our technology and experience, CD Formulation can provide technology development for the long-lasting development of protein peptide drugs and offer glycosylation modification of protein drugs to our customers.
Glycosylation is an important post-translational modification of proteins. Glycosylation of proteins can be divided into N-glycosylation and O-glycosylation depending on the way the glycan chain is connected to the peptide chain by N-acetylglucosamine (Glc-NAc) at the reducing end of the glycan chain and nitrogen atoms on some Asn side chain acylamides in the peptide chain. The sugar chains on the protein surface can affect the pharmacokinetic effects, biological activity, stability, susceptibility to proteases, water solubility, agglutination, and immunogenicity of proteins.
Glycosylation usually occurs in macromolecules such as nucleic acids, proteins and lipids, and plays an important role in biological processes such as inflammation, immune response and intracellular transport. Many therapeutic proteopeptide drugs can have improved pharmacokinetic and pharmacodynamic properties when glycosylated in vitro, and in addition, glycan modification using enzymatic or chemical conjugation methods can facilitate protein targeting to disease-affected tissues.
The glycan chains on the protein surface can influence the pharmacokinetic properties, biological activity and stability of the protein. Comparing the glycosylated and non-glycosylated forms of therapeutic proteins, on the one hand, the increased side chains on the protein drug surface improve protein stability and hinder the degradation of protein drugs by proteases, and on the other hand, they increase the molecular mass of protein drugs and reduce glomerular filtration.
Glycospectrometry assay can be performed for glycosylated drugs. After getting the theoretical sequence from the client, the sequence is first analyzed to confirm the suitable protease for the project; usually, more than two proteases are selected for enzymatic cleavage to improve the peptide coverage; different proteases are used for enzymatic cleavage to produce different peptide sequences, and the complementary relationship between the peptide sequences is used to help improve the peptide coverage for glycospectrometry analysis.
Considering the great impact of glycan structure on the half-life, efficacy and safety of antibody drugs, it is important to select highly sensitive analytical tools to characterize N-glycan heterogeneity during the development of innovative drugs and biosimilars.
Intact protein level analysis provides molecular weight information and the relative content of different glycoforms. After reducing the antibody, information on post-translational modifications such as lysine deletion, oxidation, etc. can be obtained more accurately.
Peptide map analysis is usually advantageous when the antibody contains multiple glycosylation sites. A combination of multiple proteases is used to enzymatically digest the antibody into glycopeptide fragments, which are separated and analyzed using RP-LCMS/MS.
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