A Family of Activation Associated Secreted Protein (ASP) Homologues of Cooperia Punctata

Activation-associated secreted proteins (ASP) of nematodes have been studied as potential vaccine components. In this study we report the cloning and analysis of cDNA and genomic sequences of Cooperia punctata and establish the presence of two 75% identical ASP-1 genes in C. punctata. Additional C. punctata ASP paralogues were shown to be present. Analysis of PCR products amplified from genomic DNA from a pool of worms revealed extensive sequence diversity within this family of proteins, reflecting the presence of different ASP paralogues in a single worm as well as extensive polymorphisms between different worms. ASP proteins contain a conserved region called the sperm-coating protein (SCP) domain of unknown function, which is present as a single copy in proteins from yeast and a wide range of multi-cellular organisms. Only in three nematodes has a protein composed of duplicated SCP-domains been identified. C. punctata is the first organism in which at least two such genes are found. Database searches identified similarity of the C-terminal cysteine-rich domain of ASP proteins to a nematode metallothionein motif. Cp-asp-1b was expressed in Escherichia coli and both the N-terminal and C-terminal domain were shown to be recognized by sera of C. punctata infected bovines. The description of the asp gene family of C. punctata provides the basis for more detailed studies into the extent of variation and immunological recognition of this family that may assist in rational vaccine design.