NMR Relaxation of Nuclei of Buffer as a Probe for Monitoring Protein Solutions Including Aggregation Processes
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Applied Magnetic Resonance
ORIGINAL PAPER
NMR Relaxation of Nuclei of Buffer as a Probe for Monitoring Protein Solutions Including Aggregation Processes S. O. Rabdano1 · S. S. Bystrov1 · D. A. Luzik1 · V. I. Chizhik1 Received: 26 June 2020 / Revised: 14 July 2020 © Springer-Verlag GmbH Austria, part of Springer Nature 2020
Abstract Characterization of protein solutions is of great importance for biophysical research, pharmaceutical industry, and medicine. Particularly, the monitoring of the protein aggregation is crucial at all stages of biotechnological production and in the diagnosis of dangerous diseases. The present work is focused on a study of prospects and possibilities of NMR relaxation of solvent nuclei for monitoring the state of proteins in solutions. The spin–lattice and spin–spin relaxation rates (R1 and R2) of solvent nuclei were measured in the solutions of a small globular protein, RRM2 domain of TDP-43 protein. The solvent was either H 2O- or D2O-based buffer with pH 6.5 and contained 20 mM sodium phosphate and 150 mM NaCl. The relaxation rates of the solvent 1H, 2H, 23Na, and 35Cl nuclei in solutions of soluble and aggregated RRM2 domain of TDP-43 protein were studied. The aggregation was induced by mild oxidative stress, using treatment by hydrogen peroxide. It was found that aggregation of protein could be detected using NMR relaxation of 1H nuclei. The observed CPMG dispersion for R2 rates confirms the millisecond timescale for the hydrogen exchange between water and protein sites. The correlation times and binding constants for sodium and chlorine ions were estimated using concentration dependences for relaxation rates (23Na, 35Cl). The relaxation rates of solvent nuclei are sensitive to the presence of protein in solution even at low protein concentrations, and the relaxation rates of different nuclei reflect various aspects of the state of the protein.
1 Introduction Characterization of protein solutions is of great importance for biophysical research, pharmaceutical industry, and medicine. Particularly, the monitoring of the protein aggregation is crucial at all stages of biotechnological production and
* S. O. Rabdano [email protected] 1
Saint Petersburg State University, 7/9 Universitetskaya nab., St. Petersburg 199034, Russia
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in the diagnosis of dangerous diseases. For example, the assembly of proteins to regular fibrillar structures [1–3] or inhomogeneous aggregate particles [4] is a hallmark of various neurodegenerations. Some protein aggregates perform a functional role in cells [5–7]. In biotechnology, protein aggregation sometimes used for purification of the final product, but usually lead to loss of expensive material. Most researchers who work with proteins have faced with phenomena of aggregation. The aggregates of proteins are usually defined as structures with several protein chains connected by either covalent or non-covalent interactions [8]. The protein chains are polyvalent and allow the buildup of complexes
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