Application of Natural Isotopic Abundance ¹H-¹³C- and ¹H-¹⁵N-Correlated Two-Dimensional NMR for Evaluation of the Structure of Protein Therapeutics

Methods in Enzymology
Luke W ArbogastJohn P Marino

Abstract

Methods for characterizing the higher-order structure of protein therapeutics are in great demand for establishing consistency in drug manufacturing, for detecting drug product variations resulting from modifications in the manufacturing process, and for comparing a biosimilar to an innovator reference product. In principle, solution NMR can provide a robust approach for characterization of the conformation(s) of protein therapeutics in formulation at atomic resolution. However, molecular weight limitations and the perceived need for stable isotope labeling have to date limited its practical applications in the biopharmaceutical industry. Advances in NMR magnet and console technologies, cryogenically cooled probes, and new rapid acquisition methodologies, particularly selective optimized flip-angle short transient pulse schemes and nonuniform sampling, have greatly ameliorated these limitations. Here, we describe experimental methods for the collection and analysis of 2D (1)H(N)-(15)N-amide- and (1)H-(13)C-methyl-correlated spectra applied to protein drug products at natural isotopic abundance, including representatives from the rapidly growing class of monoclonal antibody (mAb) therapeutics. Practical aspects of experimental s...Continue Reading

Citations

Feb 8, 2018·Analytical and Bioanalytical Chemistry·John E SchielKaren Phinney
Feb 13, 2018·Analytical and Bioanalytical Chemistry·Abigail TurnerJohn E Schiel
Feb 13, 2018·Analytical and Bioanalytical Chemistry·John E Schiel, Abigail Turner
Nov 30, 2018·Journal of Biomolecular NMR·Luke W ArbogastJohn P Marino
Sep 10, 2019·Journal of Magnetic Resonance·Robert G Brinson, John P Marino
Dec 15, 2020·Nature Communications·Alistair M JaggerDavid A Lomas
Dec 31, 2020·International Journal of Molecular Sciences·Yuji Tokunaga, Koh Takeuchi
Mar 5, 2021·ACS Pharmacology & Translational Science·Jack E BramhamAlexander P Golovanov
Apr 30, 2017·Journal of Pharmaceutical and Biomedical Analysis·Robert G BrinsonJohn P Marino

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