title: Further characterization of the mouse sperm surface zona-binding site with galactosyltransferase activity.
abstract: One of the mouse sperm surface binding sites for zona pellucida ligands exhibits galactosyltransferase (GT) enzyme activity. The present study was undertaken to ascertain whether the GT site behaves as a noncatalytic binding site in its physiological capacity, with no glycosylation of zona ligands, or whether glycosylation of zona ligands is an integral part of sperm-zona binding. The effects of Mn2+, the obligatory cation for GT catalysis, on enzyme activity and sperm-zona binding were examined. With uridine-5'-diphosphogalactose (UDPgal) as galactose donor, and N-acetylglucosamine (GlcNAc) as galactose acceptor, increasing concentrations of Mn2+ in the range of 0.1-10 mM increased GT enzyme activity, with half-maximal activation at 0.65 mM Mn2+ (Vmax = 20 pmol/hr/10(6) cells). In the presence of 0-2 mM Mn2+, sperm-zona binding was inhibited in a concentration-dependent manner; 50% inhibition occurred at 1.25 mM Mn2+. At this concentration, GT enzyme activity was at 65% Vmax. To determine the specificity of the GT site for glycoprotein terminal carbohydrate residues, spermatozoa were incubated with, asialo-ovine submaxillary mucin (N-acetylgalactosamine residues), asialo-, -alpha 1-acid glycoprotein (beta 1-4 galactose residues) ovalbumin (Ov; GlcNAc residues), and asialo-agalacto-/alpha 1-acid glycoprotein (AsAgAGP; GlcN-Ac residues). Only Ov and AsAgAGP acted as acceptors for galactose in the enzyme assay and inhibitors in the sperm-zona binding assay. The kinetics of the interaction of AsAgAGP with the GT site were determined: the Km was 3.6 mg/ml, with Vmax of 33 pmol/hr/10(6) cells.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Obstetrics and Gynecology, University of Pennsylvania School of Medicine, Philadelphia 19104-6080.
authors.forename: D A
authors.initials: DA
authors.lastname: Benau
authors.forename: E J
authors.initials: EJ
authors.lastname: McGuire
authors.forename: B T
authors.initials: BT
authors.lastname: Storey
mesh_terms: D000818:Animals; D001212:Asialoglycoproteins / Q000302:isolation & purification; D002462:Cell Membrane / Q000201:enzymology / Q000502:physiology; D005260:Female; D005700:Galactosyltransferases / Q000378:metabolism*; D007700:Kinetics; D007811:Lanthanum / Q000494:pharmacology; D008297:Male; D008345:Manganese / Q000494:pharmacology; D051379:Mice; D010063:Ovum / Q000502:physiology*; D013084:Sperm-Ovum Interactions / Q000187:drug effects; D013094:Spermatozoa / Q000201:enzymology* / Q000502:physiology; D015044:Zona Pellucida / Q000502:physiology*
chemical_list: D001212:Asialoglycoproteins; D008345:Manganese; D007811:Lanthanum; D005700:Galactosyltransferases