title: Conformational changes in oxidatively stressed monoclonal antibodies studied by hydrogen exchange mass spectrometry.
abstract: Oxidation of methionine residues in biopharmaceuticals is a common and often unwanted modification that frequently occurs during their manufacture and storage. It often results in a lack of stability and biological function of the product, necessitating continuous testing for the modification throughout the product shelf life. A major class of biopharmaceutical products are monoclonal antibodies (mAbs), however, techniques for their detailed structural analysis have until recently been limited. Hydrogen/deuterium exchange mass spectrometry (HXMS) has recently been successfully applied to the analysis of mAbs. Here we used HXMS to identify and localise the structural changes that occurred in a mAb (IgG1) after accelerated oxidative stress. Structural alterations in a number of segments of the Fc region were observed and these related to oxidation of methionine residues. These included a large change in the hydrogen exchange profile of residues 247-253 of the heavy chain, while smaller changes in hydrogen exchange profile were identified for peptides that contained residues in the interface of the C(H)2 and C(H)3 domains.
authors.affiliation: Department of Mass Spectrometry, LGC, Queens Road, Teddington, London, TW11 0LY, United Kingdom. william.burkitt@lgc.co.uk
authors.forename: William
authors.initials: W
authors.lastname: Burkitt
authors.forename: Paula
authors.initials: P
authors.lastname: Domann
authors.forename: Gavin
authors.initials: G
authors.lastname: O'Connor
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D003903:Deuterium / Q000737:chemistry; D041961:Deuterium Exchange Measurement / Q000379:methods*; D006859:Hydrogen / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D011487:Protein Conformation
chemical_list: D000911:Antibodies, Monoclonal; D006859:Hydrogen; D003903:Deuterium