In bovine binucleate trophoblast giant cells, pregnancy-associated glycoproteins and placental prolactin-related protein-I are conjugated to asparagine-linked N-acetylgalactosaminyl glycans.
Bovine binucleate trophoblast giant cells (BNCs) produce large amounts of PAS-positive cytoplasmic granules. After fusion of BNCs with uterine epithelial cells, the contents of these granules are released into the maternal stroma which underlies the uterine epithelium. Histochemically, the granules can be labeled with N-acetylgalactosamine-specific lectins ( Dolichos biflorus, Vicia villosa, and Wisteria floribunda agglutinins) and with Phaseolus vulgaris leucoagglutinin. In this study, we used lectin western blot analysis of proteins from fetal cotyledons to characterize the lectin binding glycoproteins. Lectin western blots showed several bands. A main band of approximately 65 kDa was identified as pregnancy-associated glycoproteins (PAGs) and a double band at 34-35 kDa as prolactin-related protein-I (PRP-I) by their crossreactivity with specific antisera. Enzymatic cleavage of N-linked glycans with peptide- N-glycanase F abolished the lectin binding to PRP and PAGs in western blots, revealing that the lectins bound to asparagine-linked glycans. The high specificity of the lectins was used for the enrichment of PRP-I and PAGs from placental cotyledons with Vicia villosa lectin affinity chromatography. The occurrence of the relatively uncommon asparagine-linked N-acetylgalactosaminyl glycans on secretory proteins of the BNCs suggests a functional role of this specific glycosylation pattern.
Klisch
Karl
K
Centre of Anatomy, University School of Medicine Hannover, Carl-Neuberg Strasse 1, 30625, Hannover, Germany. klisch.karl@mh-hannover.de
Leiser
Rudolf
R
D000116:Acetylgalactosamine / Q000737:chemistry; D000818:Animals; D001216:Asparagine / Q000737:chemistry; D015153:Blotting, Western; D002417:Cattle; D002846:Chromatography, Affinity; D005260:Female; D006023:Glycoproteins / Q000737:chemistry* / Q000302:isolation & purification; D006031:Glycosylation; D037102:Lectins; D015335:Molecular Probes; D011134:Polysaccharides / Q000737:chemistry; D011247:Pregnancy; D011257:Pregnancy Proteins / Q000737:chemistry* / Q000302:isolation & purification; D011388:Prolactin / Q000737:chemistry*; D014327:Trophoblasts / Q000737:chemistry / Q000166:cytology*
D006023:Glycoproteins; D037102:Lectins; D015335:Molecular Probes; D011134:Polysaccharides; D011257:Pregnancy Proteins; D001216:Asparagine; D011388:Prolactin; D000116:Acetylgalactosamine