title: Structural analysis of the asparagine-linked glycan units of the ZP2 and ZP3 glycoproteins from mouse zona pellucida.
abstract: Zona pellucida (ZP), the extracellular glycocalyx that surrounds the mammalian egg plasma membrane, is a relatively simple structure consisting of three to four glycoproteins. In the mouse, the ZP is composed of three glycoproteins, namely ZP1 (200 kDa), ZP2 (120 kDa), and ZP3 (83 kDa). Extensive studies in this species have resulted in the identification of primary (mZP3) and secondary (mZP2) binding sites for spermatozoa. The two zona components are highly glycosylated containing N-linked and O-linked glycan units. In an attempt to characterize N-linked glycan units, mZP2 and mZP3 were purified and the N-linked carbohydrate chains were released by exhaustive digestion with N-glycanase. The released oligosaccharides (OSs) were radiolabeled by reduction with NaB3H4 and resolved by gel filtration on a column of Bio-Gel P-4. The OSs separated into several peaks indicating the presence of a variety of N-linked glycans. Interestingly, the radioactive peaks resolved from mZP2 and mZP3 were quite different, a result suggesting qualitative and quantitative differences in the glycans. The [SH]-labeled glycans present in mZP2 and mZP3 were pooled separately and fractionated by serial lectin chromatography. Experimental evidence included in this report strongly suggests that mZP3 (but not mZP2) contains polylactosaminyl glycan with terminal, nonreducing alpha-galactosyl residues. The mZP3 glycans eluted from the immobilized lectin columns were further characterized by lectin and sizing column chromatography before or after digestion with endo-/ exo-glycohydrolases. Data revealed the presence of a variety of OSs, including poly-N-acetyllactosaminyl, bi-, tri-, and tetraantennary complex-type, and high-mannose-type glycans. Taken together, these results provide additional evidence on the complex nature of the glycan chains present on mZP glycoconjugates.
authors.affiliation: Department of Obstetrics & Gynecology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2633, USA. daulat.tulsiani@mcmail.vanderbilt.edu
authors.forename: D R
authors.initials: DR
authors.lastname: Tulsiani
mesh_terms: D000818:Animals; D001216:Asparagine / Q000737:chemistry; D002240:Carbohydrate Sequence; D002846:Chromatography, Affinity; D004527:Egg Proteins / Q000737:chemistry*; D005260:Female; D008562:Membrane Glycoproteins / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000737:chemistry; D011956:Receptors, Cell Surface; D015044:Zona Pellucida / Q000737:chemistry*; D000072216:Zona Pellucida Glycoproteins
chemical_list: D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011134:Polysaccharides; D011956:Receptors, Cell Surface; D000072216:Zona Pellucida Glycoproteins; C000606519:Zp1 protein, mouse; C000606520:Zp2 protein, mouse; C000606521:Zp3 protein, mouse; D001216:Asparagine