COMPARISON OF THE CATHEPSIN-D FROM MACKEREL (SCOMBER-AUSTRALASICUS) AND MILKFISH (CHANOS-CHANOS) MUSCLE

Muscle proteases from mackerel and milkfish were purified to electrophoretical homogeneity by concanavalin A-Sepharose and Sephadex G-100 chromatographies. Both proteases appear to be an aspartic protease, cathepsin D (EC 3.4.23.5). The molecular weights of the purified cathepsin D's from mackerel and milkfish were 51,000 and 54,000, estimated by Sephadex G-100, and 59,000 and 61,000 by SDS-PAGE, respectively. Both cathepsin D's were completely inhibited by pepstatin, but not affected by leupeptin, N-ethylmaleimide, dithiothreitol, or glutathione. Beta-Mercaptoethanol, iodoacetic acid, p-chloromercuribenzoate, phenylmethylsulfonyl fluoride, and sodium dodecyl sulfate partially or completely inhibited both cathepsin D's. Na+ and K+ partially activated the cathepsin D from milkfish. Both cathepsin D's were inhibited by Mg2+, Sr2+, Fe2+, and Hg2+, but activated by Ca2+, Co2+, Ni2+, Cu2+, Zn2+, and Cd2+. The pI and optimal temperature of the cathepsin D's from mackerel and milkfish were 5.04 and 4.91, 45-degrees, and 50-degrees-C, respectively. The temperatures for inactivating 50% activity of the cathepsin D's from mackerel and milkfish during 20 min of incubation were 53-degrees and 48-degrees-C, respectively. Both cathepsin D's had similar optimal pHs near 3. The activity of that from milkfish markedly decreased when the pH was higher than 4, and was almost completely lost at pH above 6, while that from mackerel still had at least 40% activity at pH 6.