PURIFICATION AND PROPERTIES OF TRYPSIN-LIKE ENZYMES AND A CARBOXYPEPTIDASE A FROM EUPHAUSIA SUPERBA

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Abstract
Five proteases designated as A1, A2, B, C and D were isolated from Euphausia superba by the succesive steps of ammonium sulfate fractionation, acetone precipitation, gel filtration and DEAE-Sephadex A-50 chromatography, A1, B, C and D were purified to homogeneity in disc gel electrophoresis by means of rechromatography on the DEAE-Sephadex A-50 column. Studies on substrate specificity of these enzymes revealed that A2, B, C and D were trypsin-like enzymes and A1 a carboxypeptidase A. A1, B, C and D had molecular weights of about 24,000, 24,000 28,000 and 27,000; optimal pH at 9.0, 8.0, 7.5 and 7.5–8. 0; and optimal temperature at 48, 50, 55–60 and 55°C, respectively. The activity of B, C and D were not inhibited by sulfhydryl reagents and N-α-tosyl-L-phenylalanylchloro-methyl ketone, but inhibited by reducing agents, N-α-tosyl-L-lysylchloromethyl ketone and soybean trypsin inhibitor. The activity of protease A1 was stimulated 3.5 fold by cobalt, but inhibited by 3-indolepropionate and D-phenylalanine.
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