Identification by affinity chromatography of boar sperm membrane-associated proteins bound to immobilized porcine zona pellucida. Mapping of the phosphorylethanolamine-binding region of spermadhesin AWN.
We have identified boar sperm membrane components recovered by affinity chromatography on a porcine zona pellucida affinity column. The major zona pellucida-bound proteins were spermadhesins AWN and AQN-3, the heparin-binding protein pAIF, and a homolog of the mouse milk fat globule membrane protein. All these proteins are phospholipid-binding proteins peripherally associated with the plasma membrane. Our data suggest that coating proteins tightly bound to the external lipid bilayer may act as major zona pellucida-binding molecules. Using a synthetic peptide approach we show that the regions of spermadhesin AWN comprising residues 6-12 and 104-108 possess affinity for phosphorylethanolamine. These two amino acid sequences are in close proximity in the predicted structural model for AWN, and in opposite location to its carbohydrate-recognition domain. Taken together, our data provide further evidence for the possible involvement of members of the porcine spermadhesin protein family in gamete interaction and suggest a model for the ultrastructural disposition of functional domains of spermadhesin AWN bound to the sperm surface.
Ensslin
M
M
Institut für Reproduktionsmedizin, Tierärztliche Hochschule Hannover, Germany.
Calvete
J J
JJ
Thole
H H
HH
Sierralta
W D
WD
Adermann
K
K
Sanz
L
L
Töpfer-Petersen
E
E
D000176:Acrosin / Q000037:antagonists & inhibitors*; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry*; D002846:Chromatography, Affinity; D015151:Immunoblotting; D066298:In Vitro Techniques; D008297:Male; D008562:Membrane Glycoproteins / Q000032:analysis*; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry; D010714:Phosphatidylethanolamines / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D029589:Seminal Plasma Proteins; D013094:Spermatozoa / Q000737:chemistry*; D013552:Swine; D015044:Zona Pellucida / Q000737:chemistry*
D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D010455:Peptides; D010714:Phosphatidylethanolamines; D029589:Seminal Plasma Proteins; C425042:spermadhesin; D000176:Acrosin