Hydrogen-Deuterium Exchange Dynamics of NISTmAb Measured by Small Angle Neutron Scattering

Published

November 14, 2023

Author(s)

Roisin Donnelly, Sai Pingali, Luke Heroux, Robert Brinson, Norman J. Wagner, Yun Liu

Abstract

Understanding protein dynamics and conformational stability holds great significance in biopharmaceutical research. Hydrogen−deuterium exchange (HDX) is a quantitative methodology used to examine these fundamental properties of proteins. HDX involves measuring the exchange of solvent-accessible hydrogens with deuterium, which yields valuable insights into conformational fluctuations and conformational stability. While mass spectrometry is commonly used to measure HDX on the peptide level, we explore a different approach using small-angle neutron scattering (SANS). In this work, SANS is demonstrated as a complementary and noninvasive HDX method (HDX-SANS). By assessing subtle changes in the tertiary and quaternary structure during the exchange process in deuterated buffer, along with the influence of added electrolytes on protein stability, SANS is validated as a complementary HDX technique. The HDX of a model therapeutic antibody, NISTmAb, an IgG1κ, is monitored by HDX-SANS over many hours using several different formulations, including salts from the Hofmeister series of anions, such as sodium perchlorate, sodium thiocyanate, and sodium sulfate. The impact of these formulation conditions on the thermal stability of NISTmAb is probed by differential scanning calorimetry. The more destabilizing salts led to heightened conformational dynamics in mAb solutions even at temperatures significantly below the denaturation point. HDX-SANS is demonstrated as a sensitive and noninvasive technique for quantifying HDX kinetics directly in mAb solution, providing novel information about mAb conformational fluctuations. Therefore, HDX-SANS holds promise as a potential tool for assessing protein stability in formulation.

Citation

Molecular Pharmaceutics

Volume

20

Issue

12

Pub Type

Journals

Download Paper

https://doi.org/10.1021/acs.molpharmaceut.3c00751

Local Download

Keywords

monoclonal antibody, NISTmAb, protein, SANS, HDX, dynamics

Citation

Donnelly, R. , Pingali, S. , Heroux, L. , Brinson, R. , Wagner, N. and Liu, Y. (2023), Hydrogen-Deuterium Exchange Dynamics of NISTmAb Measured by Small Angle Neutron Scattering, Molecular Pharmaceutics, [online], https://doi.org/10.1021/acs.molpharmaceut.3c00751, https://tsapps.nist.gov/publication/get_pdf.cfm?pub_id=956272 (Accessed November 20, 2024)