Diverse lectin-binding specificity of four ZP3 glycoprotein isoforms with a discrete isoelectric point in chicken egg coat.

Abstract The vertebrate egg coat corresponding to mammalian zona pellucida is a filamentous matrix composed of highly and heterogeneously glycosylated proteins designated ZP glycoproteins including ZP1 to 4, ZPD and ZPAX, and play important roles in species-specific egg–sperm interactions. Recent advance in structural biology of chicken ZP3 provided new insights into molecular mechanisms of the egg-coat function involving its carbohydrate moieties. In this study, chicken ZP3 was separated into four major and distinct isoforms with different pI in 2D-PAGE. To investigate the meanings of the ZP3 heterogeneity in egg–sperm interactions, we preliminary analyzed glycan diversity on the molecules by using lectin-staining assays. The four major ZP3 isoforms 4–7 (from acidic to basic) were recognized equally with PNA (Galβ1-3Gal N Ac), but the isoforms 5–7 were recognized dominantly with WGA ((β-Glc N Ac)n, clustered Sia), PHA-E (bi- and triantennary N -glycan containing Galβ1-4Glc N Acβ1-2Manα1-6) and RCA I (terminal Galβ1-4Glc N Ac), respectively. Despite such sugar chain diversity among the ZP3 isoforms, a partner in the egg coat, ZP1, showed specific binding to each isoform equally. Localization of ZP1 and ZP3 in the egg-coat matrix were also analyzed.