Ishidoh K, Takeda-Ezaki M, Watanabe S, Sato N, Aihara M, Imagawa K, Kikuchi M, Kominami E
Department of Biochemistry Otsuka Pharmaceutical Co. Ltd., Kawauchi-cho, Tokushima, 771-01, Japan.
J Biochem. 1999 Apr;125(4):770-9. doi: 10.1093/oxfordjournals.jbchem.a022348.
Lysosomal proteinases are translated as preproforms, transported through the Golgi apparatus as proforms, and localized in lysosomes as mature forms. In this study, we analyzed which subclass of proteinases participates in the processing of lysosomal proteinases using Bafilomycin A1, a vacuolar ATPase inhibitor. Bafilomycin A1 raises lysosomal pH resulting in the degradation of lysosomal proteinases such as cathepsins B, D, and L. Twenty-four hours after the withdrawal of Bafilomycin A1, NIH3T3 cells possess these proteinases in amounts and activities similar to those in cells cultured in DMEM and 5% BCS. In the presence of various proteinase inhibitors, procathepsin processing is disturbed by E-64-d, resulting in abnormal processing of cathepsins D and L, but not by APMSF, Pepstatin A, or CA-074. In the presence of Helicobacter pylori Vac A toxin, which prevents vesicular transport from late endosomes to lysosomes, the processing of procathepsins B and D occurs, while that of procathepsin L does not. Thus, procathepsins B and D are converted to their mature forms in late endosomes, while procathepsin L is processed to the mature form after its arrival in lysosomes by some cysteine proteinase other than cathepsin B.
溶酶体蛋白酶最初以前体形式被翻译，作为酶原形式通过高尔基体转运，并以成熟形式定位于溶酶体中。在本研究中，我们使用液泡型ATP酶抑制剂巴弗洛霉素A1分析了哪种蛋白酶亚类参与溶酶体蛋白酶的加工过程。巴弗洛霉素A1会提高溶酶体pH值，导致溶酶体蛋白酶如组织蛋白酶B、D和L的降解。在撤除巴弗洛霉素A1 24小时后，NIH3T3细胞中这些蛋白酶的含量和活性与在含有5%牛血清的DMEM中培养的细胞相似。在存在各种蛋白酶抑制剂的情况下，E-64-d会干扰组织蛋白酶原的加工过程，导致组织蛋白酶D和L的加工异常，但氨肽酶抑制剂、胃蛋白酶抑制剂A或CA-074不会产生这种影响。在存在幽门螺杆菌空泡毒素的情况下，该毒素会阻止从晚期内体到溶酶体的囊泡运输，组织蛋白酶原B和D的加工过程仍会发生，而组织蛋白酶原L则不会。因此，组织蛋白酶原B和D在晚期内体中转化为成熟形式，而组织蛋白酶原L在到达溶酶体后由除组织蛋白酶B之外的某些半胱氨酸蛋白酶加工成成熟形式。
