Mechanisms of the intracellular targeting of lysosomal cysteine proteinase

用培养的巨噬细胞研究了溶酶体半胱氨酸蛋白酶对溶酶体的细胞内靶向作用。组织蛋白酶B、H和L在高尔基体中为前酶，在溶酶体中为单链成熟酶。金属蛋白酶抑制剂和胃抑素延缓了原酶向成熟酶的转化。短脉冲追踪实验显示在转化过程中检测到中间产物，表明不同的蛋白酶在转化过程中起作用。对组织蛋白酶H和L进行了cDNA克隆和测序，并推导出酶的前前肽氨基酸序列。免疫电镜观察显示，在溶酶体中检测到成熟酶，在高尔基体中检测到原酶。前溶酶体中存在何种形式的酶，目前正在研究中。合成的蛋白酶原有30%以上在未到达溶酶体的情况下排泄到培养基中。形态学上，检测到抗肝组织蛋白酶L标记的小边，提示这些小边处于分泌途径。

Inatracellular targeting of lysosomal cysteine proteinases to lysosomes was studied with cultured macrophages. Cathepsins B, H and L were found to be as proenzymes in Golgi apparatus, and then as the single chain mature enzymes in lysosomes. Metallo-proteinase inhibitors and pepstatin retarded the conversion of proenzymes to mature enzymes. Short pulse-chase experiment showed the detection of intermediates during the conversion, suggesting that different proteinases operate in the conversion process. cDNA cloning and sequencing of cathepsin H and L were carried out and amino acid sequences of pre-propeptides of enzymes were deduced. Immunoelectron microscopic studies with antibodies specific for procathepsins showed that mature enzymes were detected in lysosomes, and that proenzymes were localized in Golgi apparatus. It is under investigation what forms of enzymes are present in prelysosomes. More than 30% of procathepsins synthesized was excreted to the medium without reaching to lysosomes. Morphologically, small vesides labeled by anticathepsin L were detected, suggesting that those are in secretory route.