A-2. リソゾーム膜の脆弱化と細胞質酵素の蛋白分解的修飾

抄録

When leupeptin, athiolproteaseinhibitorofmicrobialorigin, wasinjectedintorats, the activity of fructose-1, 6-biphosphatase [D-fructose-1, 6-biphosphate D-glyceraldehyde-3-phosphate lyase, EC 4.1. 2.13] in the liver decreased to about 70% of that in control rats. Two types of aldolase molecules isolated from the livers of control and leupeptin-treated rats, respectively, were not distinguishable on the basis of their electrophoretic mobilities, molecular weights, immunological properties or behaviours on affinity chromatography. But the specific activity of purified enzyme from livers of leupeptin-treated rats was lower than that from livers of control rat. Amino-terminal analysis of two types of aldolase indicated that decrease of aldolase activity in the liver of leupeptin treated rats is attributable to limited hydrolysis of a peptide linkage near the carboxyterminal of the peptide chain. Injection of leupeptin also caused marked increase in the activities of free lysosomal proteases, such as cathepsin A, cathepsin B, cathepsin L and cathepsin D in the cytosol fraction. After injection of leupeptin was stopped, the decreased aldolase activity and the increased cathepsin B activity in the cytosol fraction returned in parallel to the levels in control rats. A clear inverse relationship between aldolase and cathepsin B activities in the cytosol fraction was demonstrated. When insulin, which is known to stabilize the lysosomal membrane, was injected to rats simultaneously with leupeptin, both increase in free activity of cathepsin L and decrease in aldolase activity were prevented. These findings indicate that modification of aldolase may be due to action of a lysosomal protease (s). The possibility that the decrease of liver aldolase activity on administration of leupeptin to rats was produced during homogenization was excluded by showing that the aldolase activity was not changed by addition of various protease inhibitors to the homogenization medium. Enhanced sensitivity of lysosomes to osmotic shock was demonstrated in the livers of leupeptin-treated rats, suggesting that the lysosomal membrane is labilized by administration of leupeptin. This paper provides a model system to evaluate the stability of lysosomal membrane and proteolytic modification of cytosol enzymes by a released lysosomal protease (s) to the cytosol fraction.