Cathepsin L. A new proteinase from rat-liver lysosomes.
1. Cathepsin L was purified from rat liver lysosomes by cell fractionation, osmotic disruption of the lysosomes in the lysosomal mitochondrial pellet, gel filtration of the lysosomal extract and chromatography on CM-Sephadex. 2. Cathepsin L is a thiol proteinase and exists in several multiple forms visible on the disc electropherogram. By polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate its molecular weight was found to be 23000-24000. The isoelectric points of the multiple forms of cathepsin L extended from pH 5.8-6.1 ascertained by analytical isoelectric focusing. 3. Using various protein substrates, cathepsin L was found to be the most active endopeptidase from rat liver lysosomes acting at pH 6-7. In contrast to cathepsin B1, its capability of hydrolyzing N-substituted derivatives of arginine is low and it does not split esters. 4. Greatest activity is obtained close to pH 5.0 with 70-90% of maximal activity at pH 4.0 and pH 6.0 and 30-40% at pH 7.0. 5. The enzyme is strongly inhibited by leupeptin and the chloromethyl ketone of tosyl-lysine. Leupeptin acts as a pseudo-irreversible inhibitor. 6. The enzyme is stable for several months at slightly acid pH values in the presence of thiol compounds in a deep-frozen state.
D000818:Animals; D002403:Cathepsins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D002458:Cell Fractionation; D004355:Drug Stability; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007700:Kinetics; D007976:Leupeptins / Q000494:pharmacology; D008099:Liver / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology*; D046911:Macromolecular Substances; D008297:Male; D008970:Molecular Weight; D051381:Rats; D013329:Structure-Activity Relationship
D007976:Leupeptins; D046911:Macromolecular Substances; D002403:Cathepsins