Cathepsin D from the liver of the antarctic icefish Chionodraco hamatus exhibits unusual activity and stability at high temperatures1.

Cathepsin D was purified to homogeneity from the liver of Antarctic icefish by anion-exchange chromatography followed by affinity chromatography on concanavalin-A Sepharose. The purified enzyme showed a molecular mass of 40 kDa and displayed optimal activity at pH 3.0 with a synthetic chromogenic substrate. The N-terminal sequence of this proteinase was determined by automated Edman degradation and was used to design a primer for use in reverse-transcriptase polymerase chain reaction. The open reading frame of the cloned cDNA encoded an aspartic proteinase, which contained the experimentally determined N-terminal sequence. The predicted sequence (396 residues) had a high similarity with those of cathepsin D from various vertebrate sources, but was considerably different from that of nothepsin, a distinct aspartic proteinase described previously from Antarctic fish [1]. Determination of kinetic parameters for substrate hydrolysis showed that, at temperatures between 8 and 50 degrees C, the icefish cathepsin D had a higher specificity constant (kcat/Km) than human cathepsin D. The stability of both enzymes was measured at 50 degrees C and half-lives of 55 and 3 min were derived for icefish and human cathepsin D, respectively.