Synthetic nanobodies as tools to distinguish IgG Fc glycoforms

• DOI: 10.1073/pnas.2212658119
• Publication Date: 2022-11-21T20:18:11Z
• Authors: Kevin S. Kao; Aaron Gupta; Guanghui Zong; Chao Li; Isabell Kerschbaumer; Sara Borghi; Jacqueline M. Achkar; Stylianos Bournazos; Lai-Xi Wang; Jeffrey V. Ravetch
• Keywords: 0301 basic medicine; 03 medical and health sciences; SARS-CoV-2; Polysaccharides; Immunoglobulin G; Humans; COVID-19; Biological Sciences; Single-Domain Antibodies; Immunoglobulin Fc Fragments; 3. Good health

Protein glycosylation is a crucial mediator of biological functions and is tightly regulated in health and disease. However, interrogating complex protein glycoforms is challenging, as current lectin tools are limited by cross-reactivity while mass spectrometry typically requires biochemical purification and isolation of the target protein. Here, we describe a method to identify and characterize a class of nanobodies that can distinguish glycoforms without reactivity to off-target glycoproteins or glycans. We apply this technology to immunoglobulin G (IgG) Fc glycoforms and define nanobodies that specifically recognize either IgG lacking its core-fucose or IgG bearing terminal sialic acid residues. By adapting these tools to standard biochemical methods, we can clinically stratify dengue virus and SARS-CoV-2 infected individuals based on their IgG glycan profile, selectively disrupt IgG–Fcγ receptor binding both in vitro and in vivo, and interrogate the B cell receptor (BCR) glycan structure on living cells. Ultimately, we provide a strategy for the development of reagents to identify and manipulate IgG Fc glycoforms.