Analysis of sperm proteins involved in mammalian fertilization

点击翻译按钮获取中文摘要

A multiple carbohydrate recognition mechanism is working in fertilization. We have shown that several moleculesrecognizing egg zona pellucida glycoproteins are expressed on boar sperm heads.In this project, carbohydrate recognition molecules on boar sperm were investigated. When detergent-solubilized plasma membranes of boar sperm were applied to plastic wells coated with the N-acetyllactoamine-oligosaccharide probe, two molecules(70 kDa and 40 kDa) bound to the wells and binding was inhibited by addition of the oligosaccharides. Both molecules were also shown to have affinity to asialo-alpha-1-acid glycoprotein which contains N-acetyllactosamine-oligosacharides. Partial amino acid sequences of tryptic peptide fragments of the two proteins were determined. Homology search of the peptide sequences suggested that the 70 and 40 kDa molecules are members of ADAM(a disintegrin and metalloprotease protein)family protein. cDNA encoding the proteins were cloned and their nucleotide sequences were determined. Then, the recombinant proteins of porcine ADAM4 were produced by yeast and the purified proteins were shown to have affinity to integrins. The recombinant proteins of porcine ADAM5 expressed on the surface of mammalian cells were also shown to bind to asialo-fetun but not to agalacto-fetuin, suggesting that ADAM5 is a carbohydrate recognition molecule recognizing the N-acetyllactosamine structure.