title: The impact of glycosylation on monoclonal antibody conformation and stability.
abstract: Antibody glycosylation is a common post-translational modification and has a critical role in antibody effector function. The use of glycoengineering to produce antibodies with specific glycoforms may be required to achieve the desired therapeutic efficacy. However, the modified molecule could have unusual behavior during development due to the alteration of its intrinsic properties and stability. In this study, we focused on the differences between glycosylated and deglycosylated antibodies, as aglycosyl antibodies are often chosen when effector function is not desired or unimportant. We selected three human IgG1 antibodies and used PNGase F to remove their oligosaccharide chains. Although there were no detected secondary or tertiary structural changes after deglycosylation, other intrinsic properties of the antibody were altered with the removal of oligosaccharide chains in the Fc region. The apparent molecular hydrodynamic radius increased after deglycosylation based on size-exclusion chromatography analysis. Deglycosylated antibodies exhibited less thermal stability for the CH2 domain and less resistance to GdnHCl induced unfolding. Susceptibility to proteolytic cleavage demonstrated that the deglycosylated version was more susceptible to papain. An accelerated stability study revealed that deglycosylated antibodies had higher aggregation rates. These changes may impact the development of aglycosyl antibody biotherapeutics.
authors: Oceanside Pharma Technical Development, Genentech, Inc., Oceanside, CA, USA. zheng.kai@gene.com
authors: Kai
authors: K
authors: Zheng
authors: Christopher
authors: C
authors: Bantog
authors: Robert
authors: R
authors: Bayer
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D016466:CHO Cells; D002152:Calorimetry, Differential Scanning; D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D006224:Cricetinae; D006031:Glycosylation; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D013058:Mass Spectrometry; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000378:metabolism*; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase