Murine sperm-zona binding, a fucosyl residue is required for a high affinity sperm-binding ligand. A second site on sperm binds a nonfucosylated, beta-galactosyl-capped oligosaccharide.
An essential initial step in murine fertilization is the binding of acrosome-intact sperm to specific O-linked oligosaccharides on zona pellucida glycoprotein 3. While there is agreement on the primary role of O-linked glycans in this process, there is a lack of consensus on both the terminal monosaccharide(s) required for a functional sperm binding site and the corresponding protein on the sperm cell surface that recognizes this ligand. Much current debate centers on an essential role for either a terminal N-acetylglucosaminyl or, alternatively, a terminal alpha-galactosyl residue. To gain insight into the terminal saccharides required to form a functional sperm-binding ligand, dose-response curves were generated for a series of related tri- and tetrasaccharides to evaluate their relative effectiveness to competitively inhibit the in vitro binding of murine sperm to zona pellucida-enclosed eggs. A GlcNAc-capped trisaccharide, GlcNAc beta 1,4GlcNAc beta 1,4GlcNAc,was inactive (1-72 microM range). In contrast, a beta 4-galactosyl-capped trisaccharide (Gal beta 1,4GlcNAc beta 1, 4GlcNAc) and an alpha 3-galactosyl-capped trisaccharide (Gal alpha 1,3Gal beta 1,4 GlcNAc) inhibited sperm-zona binding with low or moderate affinity (ED50 = 42 microM and 5.3 microM, respectively). The addition of an alpha 3-fucosyl residue to each of these two competitive inhibitors, forming Gal beta 1,4[Fuc alpha 1,3] GlcNAc beta 1,4GlcNAc or Gal alpha 1,3Gal beta 1, 4[Fuc alpha 1,3]Glc NAc, resulted in ligands with 85- and 12-fold higher affinities for sperm, respectively (ED50 = 500 and 430 nM). Thus, the presence of a fucosyl residue appears to be obligatory for an oligosaccharide to bind sperm with high affinity. Last, mixing experiments with pairs of competitive inhibitors suggest that murine sperm-zona binding is mediated by two independent oligosaccharide-binding sites on sperm. The first (apparently high affinity) site binds both the alpha 3-galactosyl-capped trisaccharide and the two fucosylated tetrasaccharides. The second (apparently low affinity) site binds a nonfucosylated beta-galactosyl-capped trisaccharide.
Johnston
D S
DS
Department of Population Dynamics, Johns Hopkins University School of Hygiene and Public Health, Baltimore, Maryland 21205-2179, USA.
Wright
W W
WW
Shaper
J H
JH
Hokke
C H
CH
Van den Eijnden
D H
DH
Joziasse
D H
DH
D000177:Acrosome / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D004527:Egg Proteins / Q000378:metabolism*; D005260:Female; D005643:Fucose; D016256:Lewis X Antigen / Q000031:analogs & derivatives; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D017350:N-Acetylgalactosaminyltransferases / Q000235:genetics / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D013084:Sperm-Ovum Interactions; D013094:Spermatozoa / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014312:Trisaccharides / Q000378:metabolism; D015044:Zona Pellucida / Q000378:metabolism*; D000072216:Zona Pellucida Glycoproteins
D004527:Egg Proteins; D016256:Lewis X Antigen; D008562:Membrane Glycoproteins; D009844:Oligosaccharides; D011956:Receptors, Cell Surface; D014312:Trisaccharides; D000072216:Zona Pellucida Glycoproteins; C000606521:Zp3 protein, mouse; C110530:galactosyl-(1,4)-fucopyranosyl-(1,3)-N-acetylglucosamine; D005643:Fucose; D017350:N-Acetylgalactosaminyltransferases; C023220:(N-acetylneuraminyl)-galactosylglucosylceramide N-acetylgalactosaminyltransferase