Cloning, sequence and expression of rat cathepsin D.

Cathepsin D is a member of the family of aspartic proteases and is thought to play a role in the lysosomal-mediated degradation of proteins (1). We have isolated the cDNA for rat cathepsin D from a pituitary cDNA library, screened using oligonucleotide probes synthesized to the conserved active site regions of aspartic proteases (Figure 1). The cDNA contains an open reading frame which encodes for a polypeptide of 407 amino acids. The predicted amino acid sequence shows 83% and 91 % amino acid identity with human (2) and mouse (Birch, unpublished data, 3) cathepsin D. The sequence predicts two N-linked glycosylation sites at amino acid residues 134 and 258. The cDNA (1940 bp HiruHR-Xhol fragment) was subcloned into pGem3Z (Promega) and capped RNA prepared from the SP6 polymerase promoter. Incubation in a rabbit reticulocyte cell-free translation system (Promega) demonstrated that the cDNA encoded a -38000 dalton protein, which upon glycosylation increased to ~ 43500 daltons.