Purification and Some Properties of a Myofibrillar Protein-Degrading Protease, Cathepsin L, from Rabbit Skeletal Muscle1

Abstract

A newly-identified muscle protease which degraded myosin, α-actinin, and actin, and removed the Ca-sensitivity of the ATPase activity of myofibrils was purified from rabbit skeletal muscle by ammonium sulfate fractionation, Sephadex G-75 chromatography, P-cellulose chromatography, Sephadex G-75 rechromatography, and Ultrogel AcA 54 chromatography. Polyacrylamide gel electrophoresis showed that the enzyme thus obtained was almost homogeneous. The molecular weight of the enzyme was found to be 24,000 by gel filtration on Sephadex G-75. The optimum pH for the Ca-sensitivity-removing activity was pH 7.0, and that for the activity to degrade myosin was pH 4.1. The enzyme was stable at pH 4.5–6.5. It was strongly inhibited by iodoacetate, leupeptin, and antipain, but not by pepstatin or phenylmethane sulfonyl fluoride. EDTA was essential for the activity of the enzyme. The enzyme did not split α-N-benzoyl-dl-arginine-β-naphthylamide. Since these properties resemble those of cathepsin L isolated from rat liver lysosomes, the enzyme was regarded as muscle cathepsin L.