Extracellular release of the surface metalloprotease, gp63, from Leishmania and insect trypanosomatids.
Extracellular release of the surface metalloprotease, gp63, from Leishmania and insect trypanosomatids.
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利什曼原虫和昆虫锥虫的表面金属蛋白酶 gp63 的细胞外释放。
DOI：
10.1007/s00436-003-0960-0
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发表时间：
2003
影响因子：
2
通讯作者：
Dwyer,DennisM
中科院分区：
文献类型：
--
作者：
Jaffe,CharlesL;Dwyer,DennisM
在从杜氏利什曼原虫收集的废培养基中发现了蛋白酶活性。墨西哥，L.主要，以及昆虫锥虫、Crithidia luciliae 和 Leptomonas seymouri。释放的蛋白酶活性随时间线性增加，并且与前鞭毛体密度相关。在 SDS-PAGE 中，酶谱凝胶显示蛋白酶的分子量范围为 43–100 kDa。用过的培养基蛋白酶可被金属蛋白酶抑制剂、1,10-菲咯啉和 Z-Tyr-Leu-NHOH 阻断，但不会被 bestatin、leupeptin、ABESF、胃酶抑素 A、E-64 或抑肽酶阻断。针对利什曼原虫 gp63 的单克隆和/或多克隆抗体与释放的短膜虫、细单胞菌、L 反应。梅杰和L.多诺瓦尼蛋白酶。使用 gp63 特异性抗体进行的细胞表面生物素化和免疫沉淀表明，> 34% 的释放蛋白酶源自表面。针对布氏锥虫可变表面糖蛋白交叉反应决定簇 (CRD) 的抗体无法识别这种活性，这表明 gp63 并未被寄生虫磷脂酶从细胞表面裂解，而是通过替代机制释放。
Protease activity was found in spent culture medium collected fromLeishmania donovani,L. mexicana,L. major, as well as the insect trypanosomatids,Crithidia luciliaeandLeptomonas seymouri. Released protease activity increased linearly over time and was correlated to promastigote density. In SDS-PAGE, zymogram gels showed that the protease's molecular weight ranged from 43–100 kDa. Spent culture medium proteases were blocked by the metallo-protease inhibitors, 1,10-phenanthroline and Z-Tyr-Leu-NHOH, but not by bestatin, leupeptin, ABESF, pepstatin A, E-64 or aprotinin. Monoclonal and/or polyclonal antibodies to the leishmanial gp63 reacted with the releasedCrithidia,Leptomonas,L. majorandL. donovaniproteases. Cell surface biotinylation and immune precipitation using gp63-specific antibodies showed that >34% of the released protease originated from the surface. Antibodies against theTrypanosoma bruceivariable surface glycoprotein cross-reactive determinant (CRD) did not recognize this activity, suggesting that the gp63 is not cleaved from the cell surface by a parasite phospholipase, but is released by an alternative mechanism.