Profiling Formulated Monoclonal Antibodies by 1H NMR Spectroscopy.
Anal Chem. 2013 Sep 5;
Authors: Poppe L, Jordan JB, Lawson K, Jerums M, Apostol I, Schnier PD
Abstract
Nuclear magnetic resonance (NMR) is arguably the most direct methodology for characterizing the higher-order structure of proteins in solution. Structural characterization of proteins by NMR typically utilizes heteronuclear experiments. However, for formulated monoclonal antibody (mAb) therapeutics, the use of these approaches is not currently tenable due the requirements of isotope labeling, the large size of the proteins, and the restraints imposed by various formulations. Here, we present a new strategy to characterize formulated mAbs using 1H NMR. This method, based on the pulsed field gradient stimulated echo (PGSTE) experiment, facilitates the use of 1H NMR to generate highly resolved spectra of intact mAbs in their formulation buffers. This method of data acquisition, along with post-acquisition signal processing, allows the generation of structural and hydrodynamic profiles of antibodies. We demonstrate how variation of the PGSTE pulse sequence parameters allows proton relaxation rates and relative diffusion coefficients to be obtained in a simple fashion. This new methodology can be used as a robust way to compare and characterize mAb therapeutics.
PMID: 24006877 [PubMed - as supplied by publisher]
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Smith, M. J., Neel, B. G., Ikura, M....
Date: 2013-03-19
Defects in the RAS small G protein or its associated network of regulatory proteins that disrupt GTPase cycling are a major cause of cancer and developmental RASopathy disorders. Lack of robust functional assays has been a major hurdle in RAS pathway-targeted drug development. We used NMR to obtain detailed mechanistic... Read More
PNAS:
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J Mol Biol. 1998 Aug 7;281(1):61-7
Authors: Huang X, Yang X, Luft BJ, Koide S
Outer surface protein A (OspA) from the Lyme disease spirochete Borrelia burgdorferi has been a focus of vaccine development. We have identified epitopes of OspA to two monoclonal antibodies (mAbs) by comparing NMR chemical shifts of free OspA and those in Fab complexes....
Profiling Formulated Monoclonal Antibodies by 1H NMR Spectroscopy.
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