Strongyloides stercoralis, an important nematode pathogen of humans, is transmitted by contact with soil contaminated with the microscopic larvae of the parasite. We determined the cDNA sequence and deduced amino acid structure of a metallo-proteinase that is abundantly transcribed expressed by infective stage larvae of S. stercoralis. This deduced structure of the enzyme revealed a multi-domain protein that included an NH2-terminal peptidase. This peptidase consisted of a signal peptide, a pro-enzyme region, and a mature peptidase domain that included the metal ion co-ordinating motifs, HETSHALGVIH and SIMHY ("Met-turn"), characteristic of the catalytic active site of members of the metzincin superfamily of zinc metallo-endopeptidases. It was phylogenetically and structurally similar to astacin from the digestive gland of the crayfish Astacus astacus, to the HCH-1 peptidase of Caenorhabditis elegans required for hatching and migration of a post-embryonic neuroblast, and to the morphogenetically important peptidases, bone morphogenetic protein-1 (BMP-1) and Drosophila tolloid. In addition, the Strongyloides enzyme, designated strongylastacin, includes a central epidermal growth factor (EGF) domain followed by a carboxyl CUB (complement sub component C1r/C1s/embryonic sea urchin protein Uegf/bone morphogenetic protein) domain. Inspection of the dbEST database revealed the presence of at least 9 transcript clusters that are related to greater or lesser extent to strongylastacin; based on these expressed sequence tags, strongylastacin was expressed only in the infective third stage larvae, whereas other transcript clusters were expressed both in filariform and rhabditiform stages or only in the rhabditiform stage. Based on the deduced sequence, structure, and expression profile, strongylastacin is the probable candidate for the zinc-dependent metalloprotease, Ss40, known to be deployed by larvae of S. stercoralis to penetrate human skin to initiate infection.