The novel IgG-receptor FCRL5 is able to distinguish IgG2 disulfide isomers and displays increased affinity for heat-stressed IgG2

Abstract

Abstract Human Fc receptor-like 5 (FCRL5) is a novel IgG receptor that senses the whole IgG molecule, including the Fc and F(ab′)2 portions. Using surface plasmon resonance (Biacore) technology, we established that FCRL5 binds human IgG via a complex mechanism, where IgG isotype is just one determinant of the interaction. In particular, individual IgG2 samples were found to display a thousand-fold range of affinities as well as distinct kinetics, indicating that additional factors beyond the isotype affect binding. We further investigated two potential factors that may affect the interaction of IgG2 with FCRL5; structural heterogeneity related to IgG2 disulfide isoforms and charge variants mostly due to deamidation. We generated IgG2 samples enriched for either of the disulfide isoforms, IgG2-A and IgG2-B, and assessed their binding to FCRL5 using Biacore. We found that the IgG2-A isoform bound FCRL5 with up to 80-fold higher affinity than the IgG2-B isoform. IgG2 samples were also kept at 40°C for up to four weeks, a condition known to promote deamidation. We found that FCRL5 displayed higher affinity for heat-stressed IgG2, suggesting that the interaction is sensitive to deamidation of the IgG2. We conclude that FCRL5 is able to discriminate two independent changes in the IgG2 molecule, the disulfide isoform structure as well as charge variants likely related to deamidation. Of note, both conversion of disulfide isoforms and deamidation of the molecule are known to occur in vivo, on time scales comparable to the half-life of the IgG2 molecule. Therefore, our studies raise the possibility that FCRL5 discerns IgG2 variants produced in vivo in days or weeks, providing insight into the nature of the physiological FCRL5 ligand.