Therapeutic proteins products have been proven effective against various diseases. Complex post-translational modifications (PTMs) are required by most therapeutic proteins. Common PTMs include glycosylation, oxidation, deamidation, proteolysis, and so forth. These PTMS not only affect efficient secretion, drug efficacy and stability of therapeutic proteins, but also may influence safety, immunogenicity, serum clearance, pharmacokinetics, etc.
Deamidation is the process of converting asparagine and glutamine into aspartic and glutamic acids, respectively, which frequently happens during purification of natural and recombinant proteins. Deamidation might result in changes to protein hydrophobicity, charge, mass, and a reduction/loss of biological activity.
Oxidation is a major degradation pathway in protein and peptide formulation and storage, and might cause potential problem in protein production, isolation, and purification. Several amino acids are susceptible to oxidation, including methionine, cysteine, histidine, tyrosine, tryptophan and phenylalanine. Besides, oxidation of certain amino acids can cause changes of conformation, binding affinity, stability, lifespan, immunogenicity, which consequently affect the safety and efficacy of therapeutic proteins.
Since deamidation and oxidation are typical degradation for therapeutic proteins, deamidation and oxidation are critical quality attributes (CQAs) and must be presented during drug discovery.
Creative Proteomics provides a high-sensitivity deamidation and oxidation analysis service to identification and quantification of deamidation and oxidation in accordance with relevant guidelines (ICH Q6B, PharmEu and USP<1047>). Our experienced scientists and professional analytic team will deploy different approaches to help you with deamidation and oxidation analyses, and meet the requirements of every stage of drug discovery.
We Can Provide but Not Limited to:
- Asparagine Deamidation analysis
- Glutamine Deamidation analysis
- Deamidation sites identification
- Deamidation products quantification
- Oxidation analysis
- Oxidation sites identification
- Oxidation product quantification
- Glucose Unit (GU) values determination
Technology Platform of Deamidation and Oxidation Analysis Service:
Creative Proteomics provides efficient and accurate deamidation and oxidation analysis services through several techniques, including ion exchange chromatography (IEC), capillary isoelectric focusing (cIEF), hydrophobic interaction chromatography (HIC), liquid-chromatography-tandem mass spectrometry (LC-MS/MS), etc.
Both asparagine and glutamine deamidation can be rapidly analyzed by ion IEC and LC-MS/MS. And identification of oxidation sites can be determined by HIC, reverse phase-HPLC coupled with Fabricator digestion, and mixed mode size0exclusion chromatography (SEC). Creative Proteomics offers simultaneous identification and quantification of deamidation and oxidation by bottom-up LC-MS with ultrafast tryptic digestion. Pharmaceutical products will be digested into peptides by a 5 min tryptic digestion method. Then the peptides will be separated with nano-LC before subject to mass spectrometer.
Advantages of Deamidation and Oxidation Analysis Service:
- High sensitivity: deamidation and oxidation can be identified for low mass ones.
- High accuracy: deamidation and oxidation results can be consistent and accurate.
- Experienced scientists and professional analytic team: professional assistance to identify deamidation and oxidation will be provided throughout drug discovery.
- Advanced equipment: Thermo Scientific™ UltiMate™ 3000 RSLCnano System.
- Optimized analysis platform: The current technology uses automatic chromatographic instruments designed for analysis, which can reduce the scale of analysis and greatly improve the flexibility and effectiveness of analysis.
- Rapid turnaround time: 5-7 days to provide comprehensive report.
- Customized service: Optimal buffers and protocols will be customized based on your project and sample type.
Creative Proteomics's analytical scientists can provide complete analysis of deamidation and oxidation, as well as quantification of deamidation and oxidation at specific sites. Quick turnover, clear and concise written reports and protocols, and customized services to will be provided to help customers solve analytical and technical problems.
References
- Jenkins N. Modifications of therapeutic proteins: challenges and prospects. Cytotechnology. (2007) 53:121–125.
- Wang Y., Li X., Liu Y., et al. Simultaneous monitoring of oxidation, deamidation, isomerization, and glycosylation of monoclonal antibodies by liquid chromatographymass spectrometry method with ultrafast tryptic digestion. MABS. 2016, VOL. 8, NO. 8, 1477–1486.
- Nepomuceno A.I., Gibson R.J., Randall S.M., et al. Accurate identification of deamidated peptides in global proteomics using a quadrupole orbitrap mass spectrometer. J Proteome Res. 2014 February 7; 13(2).
- Riggs D.L., Silzel J.W., Lyon Y.A., et al. Analysis of glutamine deamidation: products, pathways and kinetics. Anal. Chem. 2019, 91, 13032−13038.
