Identification of the amino acid residues involved in human IgG transport into egg yolks of Japanese quail (Coturnix japonica)

In avian species, maternal immunoglobulin (Ig) Y is selectively incorporated into the yolks of maturing oocytes, although the relevance of receptor-mediated uptake is unclear. When administered to birds, several mammalian Igs, including human IgG (hIgG), are also incorporated into the yolks. In the current study, to gain insight into selective Ig transport into yolks, we intended to identify the amino acid residues critical for Ig uptake into egg yolks using alanine and glycine-scanning mutagenesis of 16 residues located along the CH2 and CH3 domains of hIgG1. Wild-type hIgG1-Fc (WT) and its mutants were synthesized, and their uptakes into the egg yolks of Japanese quail (Coturnix japonica) were determined. The triple mutation of loop MIS252–254 to GGG resulted in a 40% decrease in Fc uptake in comparison to that of the WT. Furthermore, quartet substitution of HEAL429–432 to GGGG located in an exposed loop at the CH3 domain completely abolished Fc uptake into egg yolks. Next, the residues HEAL429–432 were individually substituted with either alanine or glycine. Regardless of the glycine and alanine substitution, single mutations of H (429), E (430) and L (432) significantly reduced Fc uptake compared with WT uptake. Notably, the blood clearance rates of these mutants were equivalent to that of the WT. These results suggest that the clustered residues HEAL429–432 in the CH3 domain are important for the hIgG1 transport into the egg yolks. The sequence HEAL is conserved in chicken IgY at positions 550–553 within the CH4 domain, which might be involved in its uptake into the egg yolks by receptor-mediated endocytosis.