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

Muscle proteases from mackerel and milkfish were purified to electroph oretical homogeneity by concanavalin A-Sepharose and Sephadex G-100 ch romatographies. Both proteases appear to be an aspartic protease, cath epsin 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 S ephadex G-100, and 59,000 and 61,000 by SDS-PAGE, respectively. Both c athepsin D's were completely inhibited by pepstatin, but not affected by leupeptin, N-ethylmaleimide, dithiothreitol, or glutathione. Beta-M ercaptoethanol, iodoacetic acid, p-chloromercuribenzoate, phenylmethyl sulfonyl fluoride, and sodium dodecyl sulfate partially or completely inhibited both cathepsin D's. Na+ and K+ partially activated the cathe psin D from milkfish. Both cathepsin D's were inhibited by Mg2+, Sr2+, Fe2+, and Hg2+, but activated by Ca2+, Co2+, Ni2+, Cu2+, Zn2+, and Cd 2+. 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, respect ively. The temperatures for inactivating 50% activity of the cathepsin D's from mackerel and milkfish during 20 min of incubation were 53-de grees and 48-degrees-C, respectively. Both cathepsin D's had similar o ptimal pHs near 3. The activity of that from milkfish markedly decreas ed 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.