242
J. J. CALVETE and L. SANZ
concept that sperm-egg binding involves complementary molecules was postulated more than 80 years ago (Lillie 19l3), our understanding of the molecular
basis of fertilisation is still in its infancy.
In the pig, sperm-binding activity resides in the 55 kDa ZPa (ZPB) glycoprotein
(Yurewicz et al. 1991). However, conflicting results have been reported regarding
the identity of the glycans possessing the sperm-zona pellucida binding capability, e.g. O-linked (Yurewicz et al. 1991) versus N-linked tri- and/or tetraantennary
(Kudo et al. 1998) carbohydrate structures. On the sperm side, both integral
plasma membrane proteins (APz, zonadhesin) and peripherally associated proteins (spermadhesins, P47) have been described as potential components of the
primary gamete recognition and/or signalling sperm machinery (Ensslin et al.
1998). Among the variety of sperm proteins known to possess zona pellucida
glycoprotein- and carbohydrate-binding capabilities, spermadhesins emerge as a
novel protein family of animal lectins.
2
Spermadhesins: An Overview
Spermadhesins are a group of homologous polypeptides (Fig. 17.1), which represent over 90 % of the total boar seminal plasma proteins and coat the sperm surface at ejaculation (reviewed by Calvete et al. 1995a, 1996a; Topfer-Petersen and
Calvete, 1996; Topfer-Petersen et al. 1998). Although the major biological source
of porcine spermadhesins is the secretion of the seminal vesicle epithelium,
where the concentration of various spermadhesins (AQN-1, AQN-3, AWN, PSP-I,
PSP-II) ranges from 0.6-7 mg/ml, spermadhesin AWN is also synthesised by the
tubuli recti and rete testis. It is the only member of its protein family found on the
surface of porcine epididymal spermatozoa. About 6 million AWN molecules
have been quantitated on the surface of a single boar epididymal spermatozoon.
After ejaculation, 12-60 million of each spermadhesin AQN-1, PSP-I, AQN-3,
and AWN become coated on the apical third of the sperm acrosomal cap. Most of
this material, however, is released during in vitro capacitating conditions, and
the level of individual spermadhesins drops to 5-6 million molecules per spermatozoon. This suggests that the loosely attached spermadhesin population may
serve as decapacitation or acrosome-stabilizing factors, which protect the acrosomal membrane from premature acrosome reactions.
Increasing evidence indicates that the population of spermadhesin molecules
tightly bound to the surface of capacitated sperm may serve as primary zona
pellucida-binding molecules. Thus, spermadhesins AQN-1, AQN-3, and AWN
display cation-independent binding affinity (Kd in the 11M range) for both zona
pellucida glycoproteins and ~-galactosides in Gal~(l-3)-GalNAc and Gal~(l-4)GlcNAc sequences of mono- (O-linked) and trianntenary (N-linked) oligo sacharide structures of standard glycoproteins (DostaIova et al. 1995; Calvete et al.
1996b). Interestingly, the smallest O-linked -type, intact zona pellucida -derived,
synthetic oligosaccharide possessing sperm-zona pellucida binding inhibitory
activity has the structure Gal~(l-4)[HSOr61GlcNAc~(l-3)-Gal~(l-4)-GlcNAc~
(l-3)-Gal~(l-3)-GalNAc-ol (Spijker et al. 1996). On the other hand, Kudo et al.
J. J. CALVETE and L. SANZ
concept that sperm-egg binding involves complementary molecules was postulated more than 80 years ago (Lillie 19l3), our understanding of the molecular
basis of fertilisation is still in its infancy.
In the pig, sperm-binding activity resides in the 55 kDa ZPa (ZPB) glycoprotein
(Yurewicz et al. 1991). However, conflicting results have been reported regarding
the identity of the glycans possessing the sperm-zona pellucida binding capability, e.g. O-linked (Yurewicz et al. 1991) versus N-linked tri- and/or tetraantennary
(Kudo et al. 1998) carbohydrate structures. On the sperm side, both integral
plasma membrane proteins (APz, zonadhesin) and peripherally associated proteins (spermadhesins, P47) have been described as potential components of the
primary gamete recognition and/or signalling sperm machinery (Ensslin et al.
1998). Among the variety of sperm proteins known to possess zona pellucida
glycoprotein- and carbohydrate-binding capabilities, spermadhesins emerge as a
novel protein family of animal lectins.
2
Spermadhesins: An Overview
Spermadhesins are a group of homologous polypeptides (Fig. 17.1), which represent over 90 % of the total boar seminal plasma proteins and coat the sperm surface at ejaculation (reviewed by Calvete et al. 1995a, 1996a; Topfer-Petersen and
Calvete, 1996; Topfer-Petersen et al. 1998). Although the major biological source
of porcine spermadhesins is the secretion of the seminal vesicle epithelium,
where the concentration of various spermadhesins (AQN-1, AQN-3, AWN, PSP-I,
PSP-II) ranges from 0.6-7 mg/ml, spermadhesin AWN is also synthesised by the
tubuli recti and rete testis. It is the only member of its protein family found on the
surface of porcine epididymal spermatozoa. About 6 million AWN molecules
have been quantitated on the surface of a single boar epididymal spermatozoon.
After ejaculation, 12-60 million of each spermadhesin AQN-1, PSP-I, AQN-3,
and AWN become coated on the apical third of the sperm acrosomal cap. Most of
this material, however, is released during in vitro capacitating conditions, and
the level of individual spermadhesins drops to 5-6 million molecules per spermatozoon. This suggests that the loosely attached spermadhesin population may
serve as decapacitation or acrosome-stabilizing factors, which protect the acrosomal membrane from premature acrosome reactions.
Increasing evidence indicates that the population of spermadhesin molecules
tightly bound to the surface of capacitated sperm may serve as primary zona
pellucida-binding molecules. Thus, spermadhesins AQN-1, AQN-3, and AWN
display cation-independent binding affinity (Kd in the 11M range) for both zona
pellucida glycoproteins and ~-galactosides in Gal~(l-3)-GalNAc and Gal~(l-4)GlcNAc sequences of mono- (O-linked) and trianntenary (N-linked) oligo sacharide structures of standard glycoproteins (DostaIova et al. 1995; Calvete et al.
1996b). Interestingly, the smallest O-linked -type, intact zona pellucida -derived,
synthetic oligosaccharide possessing sperm-zona pellucida binding inhibitory
activity has the structure Gal~(l-4)[HSOr61GlcNAc~(l-3)-Gal~(l-4)-GlcNAc~
(l-3)-Gal~(l-3)-GalNAc-ol (Spijker et al. 1996). On the other hand, Kudo et al.