Primary, Secondary, and Quaternary Structure Characterization of IgG1, IgG2, IgG3, and IgG4 Fc regions

Abstract

Given the vital role that antibodies play in immune health and the expanding applications of monoclonal antibodies in therapeutics, there is a growing need for their structural analysis. As the intricacies of protein function are linked to their higher order structures, acquiring comprehensive information on all levels of protein structure of monoclonal antibodies would lend insight to their mechanisms. While this information is readily available for crystallized antibodies, such data is of greater therapeutic relevance in an aqueous environment. In this study, we employed Microfluidic Modulation Spectroscopy (MMS) to analyze the secondary structure of the Fc regions of IgG1, IgG2, IgG3, and IgG4 in formulation conditions. We found that these IgG subclasses exhibit structurally similar yet distinguishable characteristics. They shared high beta-sheet content and low alpha-helix and unordered content, with subtle differences in relative composition. Furthermore, our findings indicate that IgG1 and IgG2 exhibit the highest degree of structural similarity, while IgG3 and IgG4 show the greatest structural difference. This study underscores the potential of MMS as an essential tool to analyze the structure of IgG and other antibodies in formulation conditions, highlighting the role this technology can play within the drug development process.