title: Deciphering the Biophysical Effects of Oxidizing Sulfur-Containing Amino Acids in Interferon-beta-1a using MS and HDX-MS.
abstract: Introduction of a chemical change to one or more amino acids in a protein's polypeptide chain can result in various effects on its higher-order structure (HOS) and biophysical behavior (or properties). These effects range from no detectable change to significant structural or conformational alteration that can greatly affect the protein's biophysical properties and its resulting biological function. The ability to reliably detect the absence or presence of such changes is essential to understanding the structure-function relationship in a protein and in the successful commercial development of protein-based drugs (biopharmaceuticals). In this paper, we focus our attention on the latter by specifically elucidating the impact of oxidation on the HOS, structural dynamics, and biophysical properties of interferon beta-1a (IFNβ-1a). Oxidation is a common biochemical modification that occurs in many biopharmaceuticals, specifically in two naturally-occurring sulfur-containing amino acids, methionine and cysteine. To carry out this work, we used combinations of hydrogen peroxide and pH to differentially oxidize IFNβ-1a (to focus on only methionine oxidation versus methionine and cysteine oxidation). We then employed several analytical and biophysical techniques to acquire information about the differential impact of these two oxidation scenarios on IFNβ-1a. In particular, the use of MS-based techniques, especially HDX-MS, play a dominant role in revealing the differential effects. Graphical Abstract ᅟ.
authors.affiliation: Technical Development, Biogen, 225 Binney St., Cambridge, MA, 02142, USA.|Codiak Biosciences, 500 Technology Square, Cambridge, MA, 02139, USA.
authors.forename: Damian J
authors.initials: DJ
authors.lastname: Houde
authors.affiliation: Technical Development, Biogen, 225 Binney St., Cambridge, MA, 02142, USA. george.bou-assaf@biogen.com.
authors.forename: George M
authors.initials: GM
authors.lastname: Bou-Assaf
authors.affiliation: Technical Development, Biogen, 225 Binney St., Cambridge, MA, 02142, USA.
authors.forename: Steven A
authors.initials: SA
authors.lastname: Berkowitz
mesh_terms: D000276:Adjuvants, Immunologic / Q000737:chemistry*; D000818:Animals; D000998:Antiviral Agents / Q000737:chemistry*; D016466:CHO Cells; D002851:Chromatography, High Pressure Liquid; D003412:Cricetulus; D003545:Cysteine / Q000737:chemistry*; D006801:Humans; D006861:Hydrogen Peroxide / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D000068556:Interferon beta-1a / Q000737:chemistry*; D013058:Mass Spectrometry; D008715:Methionine / Q000737:chemistry*; D008958:Models, Molecular; D010084:Oxidation-Reduction; D010449:Peptide Mapping; D011487:Protein Conformation
chemical_list: D000276:Adjuvants, Immunologic; D000998:Antiviral Agents; D008715:Methionine; D006861:Hydrogen Peroxide; D003545:Cysteine; D000068556:Interferon beta-1a
keywords: Biophysical properties; Conformation; Conformational dynamics; Higher-order structure; Methionine and cysteine oxidation