Isolation and characterisation of two sperm membrane proteins recognised by sperm-associated antibodies in infertile men

Antisperm antibodies eluted from the surface of spermatozoa obtained from i nfertile men recognised several common epitopes. We tested whether these ep itopes were relevant to fertility by isolating the immunodominant 37/36 and 19/18 protein zones. These protein zones were cut out of preparative slab gets and electro-eluted. The isolated proteins, P36 and P18, were used for biochemical characterisation and to produce specific antibodies in rabbits. The specific reactivity of P36 and P18 with WGA and AAL lectins, respectiv ely, indicated the presence of lactosaminyl structures with sialic acid ter mini in P36 and of fucosylated residues in P18. Isoelectric focusing showed that the two proteins consist of several polypeptides. Some of these polyp eptides were recognised by both human and rabbit antibodies: the pi of thes e epitopes was around 5.5 for P36 and 8.3-10.3 for P18. Rabbit antibodies d etected the corresponding proteins on the sperm heads of methanol-fixed and of live acrosome-reacted spermatozoa. Anti-P36 antibodies bound mainly to the equatorial segment. They reduced the binding and, consequently, the pen etration of zona-free hamster oocytes by human spermatozoa. Anti-P18 antibo dies gave more diffuse staining of the acrosomal and postacrosomal regions and reduced sperm-oocyte penetration without a significant effect on sperm binding. These results suggest that P36 and P18 antigens located in differe nt compartments of the sperm head may participate in the sperm-oolemma inte raction. We are currently investigating the physiological role of these ant igens by sequencing the proteins isolated from the gels. (C) 2000 Wiley-Lis s, Inc.