Isolation and characterization of a cDNA clone encoding a thiol proteinaseof Sarcocystis muris cyst mero' zoites (Apicomplexa)

A cDNA library of Sarcocystis muris cyst merozoites was screened using a di goxigenin-labeled probe. This probe was derived from a 504-bp polymerase-ch ain-reaction fragment representing part of a thiol proteinase. Several cDNA clones were isolated, one of which (PH08) consists of a nucleotide sequenc e of 1694 bp and encodes the complete prepropolypeptide of a cathepsin L-li ke proteinase. PH08 contains an open reading frame of 394 amino acid (aa) r esidues with a 46-residue signal sequence, which is followed by a 129-resid ue propeptide and 219 aa residues of the mature enzyme. Two potential glyco sylation sites and a putative polyadenylation signal were also identified. The occurrence of the highly conserved interspersed ERFNIN aa motif, not fo und in cathepsin B-like proteinases, suggests the classification of the enz yme as a cathepsin L-like proteinase. Results worked out in this study will enable production of the recombinant thiol proteinase of S. muris cyst mer ozoites necessary for study of the substrate specificity as well as other b iochemical parameters of this enzyme.