Characterization of Pepstatin Insensitive Protease in Mackerel White Muscle

Abstract

The purified mackerel white muscle pepstatin insensitive protease was characterized. The enzyme had a pH optimum of 4.0 and a temperature optimum at 45°C when hemoglobin was used as substrate. The enzyme was strongly inactivated by leupeptin, PCMB, Hg²⁺ and Cue²⁺. The enzyme inactivated by PCMB and Hg²⁺ was activated again by addition of dithiothreitol, 2-mercaptoethanol and cysteine. Rabbit muscle aldolase was inactivated by the enzyme. The enzyme hydrolyzed Z-Phe-Arg-methylcoumarylamide much more than Bz-Phe-Val-Arg-methyl-coumarylamide. The molecular weight of the enzyme was estimated to be 48, 000 by gel filtration, and the isoelectric point was 4.2. It is suggested that the pepstatin insensitive protease belongs to a cysteine proteinase and shows some similarities to cathepsins L and S.