title: Post-translational modifications differentially affect IgG1 conformation and receptor binding.
abstract: Post-translational modifications (PTMs) can have profound effects on protein structure and protein dynamics and thereby can influence protein function. To understand and connect PTM-induced functional differences with any resulting conformational changes, the conformational changes must be detected and localized to specific parts of the protein. We illustrate these principles here with a study of the functional and conformational changes that accompany modifications to a monoclonal immunoglobulin gamma1 (IgG1) antibody. IgG1s are large and heterogeneous proteins capable of incorporating a multiplicity of PTMs both in vivo and in vitro. For many IgG1s, these PTMs can play a critical role in affecting conformation, biological function, and the ability of the antibody to initiate a potential adverse biological response. We investigated the impact of differential galactosylation, methionine oxidation, and fucosylation on solution conformation using hydrogen/deuterium exchange mass spectrometry and probed the effects of IgG1 binding to the FcgammaRIIIa receptor. The results showed that methionine oxidation and galactosylation both impact IgG1 conformation, whereas fucosylation appears to have little or no impact to the conformation. FcgammaRIIIa binding was strongly influenced by both the glycan structure/composition (namely galactose and fucose) and conformational changes that were induced by some of the modifications.
authors.affiliation: Biogen Idec, Inc., Cambridge, Massachusetts 02142, USA.
authors.forename: Damian
authors.initials: D
authors.lastname: Houde
authors.forename: Yucai
authors.initials: Y
authors.lastname: Peng
authors.forename: Steven A
authors.initials: SA
authors.lastname: Berkowitz
authors.forename: John R
authors.initials: JR
authors.lastname: Engen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003903:Deuterium; D006031:Glycosylation; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry* / Q000378:metabolism*; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011522:Protons; D011961:Receptors, Fc / Q000378:metabolism*
chemical_list: D007074:Immunoglobulin G; D050505:Mutant Proteins; D011522:Protons; D011961:Receptors, Fc; D003903:Deuterium