Structure-function relationships of human SIgA glycosylation

Wider Research Context: Secretory IgA (SIgA) has emerged as a promising candidate for therapeutic interventions owing to its unique structural features including heightened pathogen neutralization efficacy, anti-inflammatory characteristics, and enhanced stability in mucosal secretions. Its interaction with commensal bacteria and mucosal components presents an innovative avenue for novel therapeutic applications. A pivotal determinant of SIgA's unique properties is its extensive glycosylation, yet the glycosylation profile across different tissues and its biological significance remains largely unexplored. Hypotheses: We hypothesize that specific glycan modifications on SIgA are functionally crucial for interactions with components of the mucosa, commensal bacteria and host cell receptors. We aim to fill the knowledge gap on human SIgA glycosylation through a comprehensive analysis of SIgA glycan profiles in different human tissues and subsequent glyco-engineering approaches on recombinant SIgA to elucidate structure-function relationships of distinct glycoforms. Approach: We will elucidate site-specific glycosylation profiles on human SIgA isolated from different tissues, based on which we will employ glyco-engineering tools in the plant-based transient production platform Nicotiana benthamiana a proven and highly suitable system for efficiently producing this intricate and multimeric protein with tailored N- and O-glycans. Generated recombinant SIgA glycoforms will be subjected to extensive biochemical and biophysical characterization as well as in vitro and cell-based binding assays, activation assays and proteolytic stability assessments in mucosal fluids to shed light on the implication of SIgA glycosylation for mucosal immunity and therapeutic development. Innovation: This project innovatively leverages the plant-based transient production platform in Nicotiana benthamiana for glycoengineering, enabling the production of fully assembled and functional SIgA with tailored N- and O-glycans. This approach distinguishes itself from previous studies that relied on heterogeneous glycosylation profiles. The capability to produce large amounts of recombinant human SIgA with homogenous glycoforms allows for unprecedented insights into the structure-function relationships of SIgA, advancing its potential for therapeutic applications. Primary Researchers Involved: Kathrin Göritzer, based at BOKU Vienna as a postdoctoral research fellow (FWF Schrödinger fellowship) has expertise in protein and glyco-engineering of therapeutic antibodies, and has undergone extensive training in immunology. Julian Ma, based at St. George’s University of London is an internationally renowned plant biotechnologist and immunologist.