The post-translational addition of a functional group covalently bonded to the protein are common to biologic drug substances, which can affect the structure, stability, function, and protein interaction. N-acetylation is one of the post-translational modifications (PTMs), which append an acetyl group to the N-terminal amino group. N-acetylation degree might alter the charge, hydrophobicity, lifespan, folding characteristics and binding properties, and polymorphism in drug metabolism. To confirm protein identity, ensure product quality and consistent manufacturing process, and avoid a potentially harmful side effect and low efficacy in the patients, N-acetylation analysis is an important biochemical technique during the processes of drug discovery. It can be used to analyze the N-terminal modification of biological agents or target proteins.
According to ICH Q6B guideline, Creative Proteomics will perform a high-quality N-acetylation analysis services to identify N-acetylation, quantitatively analyze N-acetylation, and have better understanding of the analytical characterization of structure, comparability, stability, pharmacogenetics, and so forth. The N-acetylation analysis service can be provided according to PharmEu and USP<1047>.
We Can Provide but Not Limited to:
- N-acetylation analysis
- N-acetylation degree analysis
- Acetylation site analysis
- Quantitative analysis of N-acetylation
- Acetylated drug metabolism analysis
- Acetylated drug-protein interaction analysis
- Acetylated drug Pharmacokinetic/Pharmacodynamics (PK/PD) analysis
- Acetylated drug metabolomics analysis
Technology Platform of N-acetylation Analysis Service:
Creative Proteomics provides efficient and accurate N-acetylation analysis services in different samples and in both eukaryotic and prokaryotic organisms through High Performance Liquid Chromatography (HPLC), Mass Spectrometry (MS) analysis.
Proteins are extracted from lysed, digested by detergent-free trypsin, and then the complex mixture of peptides is separated by multi-dimensional chromatography after removal of N-Pyroglutamate. To enrich the N-acetylated peptides, the combined fractional diagonal chromatography (COFRADIC) method, or the strong cation exchange chromatography (SCX) method, or CNBr-activated sepharose resin will be employed depending on your special requirement for your project. CNBr-activated sepharose resin is simpler than previous methods, and the ratio of digest to rein will not influence the efficiency. Then enriched N-acetylated peptides are subjected to mass spectrometric analysis. To get better yield to N-acetylation identification, a MS data acquisition method, singly charged ion inclusion (SCII), will be conducted to allow the singly charged precursor.
Advantages of N-acetylation Analysis Service:
- High sensitivity: N-acetylation can be identified for low mass ones.
- Advanced equipment: High-performance liquid chromatography LC-20AT, SHIMADZU and the Thermo Scientific™ Q Exactive™ Hybrid Quadrupole-Orbitrap Mass Spectrometer.
- 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 confirmation of N-acetylation of target proteins, samples, or unknown proteins, as well as quantitative information on N-acetylation degree. 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
- Protein Acetylation Methods and Protocols (2013). Hake S.B., Janzen C.J. (Eds). Hatfield, Hertfordshire (UK): Humana Press.
- Rioux N., Mitchell L.H., Tiller P., et al., Structural and kinetic characterization of a novel N-acetylated aliphatic amine metabolite of the PRMT inhibitor, EPZ011652. Drug Metab Dispos, 43:936–943, July 2015.
- Dadou S.M., El-Barghouthi M.I., Alabdallah S.K., et al., Effect of Protonation State and N-Acetylation of Chitosan on Its Interaction with Xanthan Gum: A Molecular Dynamics Simulation Study. Marine Drugs, 2017, 15, 298.
- Ree R., Varland S., and Arnesen T., Spotlight on protein N-terminal acetylation. Experimental & Molecular Medicine, (2018) 50:90.
