The role of Ce{open}2, Ce{open}3, and Ce{open}4 domains in human and canine IgE and their contribution to Fce{open}RIα interaction

Abstract

The Ce{open}2 and Ce{open}4 domains are considered as scaffolds, allowing Ce{open}3 domains to assume an appropriate orientation to interact with Fce{open}RI (Wurzburg and Jardetzky, 2002; Hunter et al., 2008). Human/canine IgE chimeric antibodies were expressed to assess the nature of the contribution of Ce{open}2 and Ce{open}4 domains to bind to and induce target cell degranulation via Fce{open}RIα. Our results indicate that for (1) Ce{open}3 domains in IgE of canine and human origin are the only necessary region for binding to Fce{open}RIα. (2) The interaction of canine IgE with human sFce{open}RIα is significantly enhanced by contributions from both Ce{open}2 and Ce{open}4 domains of dog origin. (3) The canine/human IgE chimeric antibody construct rapidly dissociates from its the receptor when the canine Ce{open}2 and Ce{open}4 domains are replaced by the homologous human Fc domains which do not confer a conformation on the Ce{open}3 domain to facilitate stable interaction with canine Fce{open}RIα. Kinetic constants for the binding of this chimera to the soluble extracellular domain of the receptor indicate an approximate 120-fold decrease in the affinity for canine sFce{open}RIα (ka=5.30×102M-1s-1) and a 330-fold increase in the dissociation from canine sFce{open}RIα (KD=6.9×10-6M-1), compared to the wild type IgE kinetic constants (Ka=6.30×104M-1s-1; KD=2.1×10-8M-1). Although canine IgE does engage human Fce{open}RIα, canine Ce{open}2 and Ce{open}4 do not contribute to the high-affinity of interaction with human Fce{open}RIα. Upon replacement of human Ce{open}2 and Ce{open}4 domain by the canine homologues, human IgE Ce{open}3 only retains a low affinity for the human receptor, which shows that Ce{open}2 and Ce{open}4 domains in human IgE Fc contribute significantly to the interaction with its cognate receptor. © 2013 Elsevier Ltd.