Cold Denaturation of Proteins in the Absence of Solvent: Implications for Protein Storage**

Emma L. Norgate; Rosie Upton; Kjetil Hansen; Bruno Bellina; C. Brookes; Argyris Politis; Perdita E. Barran

doi:10.1002/anie.202115047

Original language	English
Article number	e202115047
Journal	Angewandte Chemie - International Edition
Volume	61
Issue number	25
DOIs	https://doi.org/10.1002/anie.202115047
Accepted/In press	2022
Published	20 Jun 2022
Additional links	Link to publication in Scopus

Bibliographical note

Funding Information: This work has been funded by a Research Impact Scholarship funded by alumni of the University of Manchester and Bristol-Myers Squibb for a CASE studentship awarded to E.N., a BBSRC Industrial Case Studentship awarded to R.U. in collaboration with Covance Laboratories and by the Leverhulme Trust (RPG-2019-178). Instrumentation was supported by Waters Corp. MS Vision, EPSRC (EP/T019328/1, GR/S77639/01) and BBSRC (BB/L01646X/1 BB/L015048/1 BB/L002655/1) awards and a lot of hard work and sweat. Funding Information: This work has been funded by a Research Impact Scholarship funded by alumni of the University of Manchester and Bristol‐Myers Squibb for a CASE studentship awarded to E.N., a BBSRC Industrial Case Studentship awarded to R.U. in collaboration with Covance Laboratories and by the Leverhulme Trust (RPG‐2019‐178). Instrumentation was supported by Waters Corp. MS Vision, EPSRC (EP/T019328/1, GR/S77639/01) and BBSRC (BB/L01646X/1 BB/L015048/1 BB/L002655/1) awards and a lot of hard work and sweat. Publisher Copyright: © 2022 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.

Abstract

The effect of temperature on the stability of proteins is well explored above 298 K, but harder to track experimentally below 273 K. Variable-temperature ion mobility mass spectrometry (VT IM-MS) allows us to measure the structure of molecules at sub-ambient temperatures. Here we monitor conformational changes that occur to two isotypes of monoclonal antibodies (mAbs) on cooling by measuring their collision cross sections (CCS) at discrete drift gas temperatures from 295 to 160 K. The CCS at 250 K is larger than predicted from collisional theory and experimental data at 295 K. This restructure is attributed to change in the strength of stabilizing intermolecular interactions. Below 250 K the CCS of the mAbs increases in line with prediction implying no rearrangement. Comparing data from isotypes suggest disulfide bridging influences thermal structural rearrangement. These findings indicate that in vacuo deep-freezing minimizes denaturation and maintains the native fold and VT IM-MS measurements at sub ambient temperatures provide new insights to the phenomenon of cold denaturation.

King's College London

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