Yamaguchi N, Kawai K
Cancer Res. 1986 Oct;46(10):5353-9.
Active acid protease was found in serum-free spent medium of human pancreatic carcinoma cell line HPC-YT. These cells have been maintained for over 215 generations in a serum-free, chemically defined medium. Acid protease was partially purified about 3000-fold by Mono Q ion-exchange chromatography, pepstatin-aminohexyl-Sepharose affinity chromatography, hydroxylapatite affinity chromatography, lectin affinity chromatographies, and gel filtration on TSK G3000SW. The molecular weight of the purified enzyme was estimated to be 63,000. An acid protease isolated from normal pancreas has two forms with molecular weights of 11,000 and 32,000. Isoelectric focusing of acid protease derived from HPC-YT revealed five isotypes, whereas the enzyme from normal pancreas showed only three bands. The optimal pH using bovine hemoglobin as substrate was 3.0 for both the normal and the HPC-YT-derived acid proteases. The HPC-YT enzyme was more heat stable than the enzymes of normal pancreas. Sulfhydryl compounds and metal ions had no apparent effects on this enzyme; however, pepstatin strongly inhibited it. We suggest that acid protease secreted from cancer cells may play a role in the destruction of the surrounding tissue matrix.
在人胰腺癌细胞系HPC-YT的无血清培养液中发现了活性酸性蛋白酶。这些细胞已在无血清、化学成分明确的培养基中传代培养超过215代。酸性蛋白酶通过Mono Q离子交换色谱、胃蛋白酶抑制剂-氨基己基-琼脂糖亲和色谱、羟基磷灰石亲和色谱、凝集素亲和色谱以及TSK G3000SW凝胶过滤进行部分纯化，纯化倍数约为3000倍。纯化酶的分子量估计为63,000。从正常胰腺分离的酸性蛋白酶有两种分子量形式，分别为11,000和32,000。对源自HPC-YT的酸性蛋白酶进行等电聚焦显示有五种同工型，而来自正常胰腺的酶仅显示三条带。以牛血红蛋白为底物时，正常和源自HPC-YT的酸性蛋白酶的最适pH均为3.0。源自HPC-YT的酶比正常胰腺的酶更耐热。巯基化合物和金属离子对该酶无明显影响；然而，胃蛋白酶抑制剂对其有强烈抑制作用。我们认为癌细胞分泌的酸性蛋白酶可能在周围组织基质的破坏中起作用。
