Anderson S A, Mukkada A J
Department of Biological Sciences, University of Cincinnati, OH 45221.
Biochim Biophys Acta. 1994 Oct 12;1195(1):71-80. doi: 10.1016/0005-2736(94)90011-6.
An ATPase on the plasma membrane of Leishmania donovani has been characterized. An antiserum, generated against ATPase active bands from native gels, was specific for a 105 kDa protein in promastigotes. However, in plasma membrane preparations a 70 kDa protein is also recognized, suggesting proteolysis of the intact 105 kDa protein or the presence of a second similar ATPase. [gamma-32P]ATP phosphorylates two proteins (105 kDa and 70 kDa) in promastigotes and plasma membranes. Both proteins form a transient phosphorylated intermediate, characteristic of a P-type ATPase. Immunostaining of permeabilized parasites shows diffuse staining of the surface of promastigotes and amastigotes, which is consistent with a plasma membrane protein. The antiserum immunoprecipitates a 70 kDa [14C]DCCD binding protein from whole cells and plasma membranes of promastigotes. Furthermore, the antiserum immunoprecipitates a 105 kDa and 70 kDa protein which can be subsequently phosphorylated. These results indicate the presence of a 105 kDa P-type ATPase on the L. donovani plasma membrane which is similar to the mammalian and fungal cation pumps.
杜氏利什曼原虫质膜上的一种ATP酶已得到鉴定。针对天然凝胶中ATP酶活性条带产生的抗血清，对前鞭毛体中的一种105 kDa蛋白具有特异性。然而，在质膜制剂中也能识别出一种70 kDa的蛋白，这表明完整的105 kDa蛋白发生了蛋白水解，或者存在第二种类似的ATP酶。[γ-32P]ATP使前鞭毛体和质膜中的两种蛋白（105 kDa和70 kDa）发生磷酸化。这两种蛋白都形成了一种瞬时磷酸化中间体，这是P型ATP酶的特征。对通透化寄生虫的免疫染色显示，前鞭毛体和无鞭毛体表面呈弥漫性染色，这与质膜蛋白一致。该抗血清从前鞭毛体的全细胞和质膜中免疫沉淀出一种70 kDa的[14C]DCCD结合蛋白。此外，该抗血清免疫沉淀出一种105 kDa和70 kDa的蛋白，随后这两种蛋白可被磷酸化。这些结果表明，杜氏利什曼原虫质膜上存在一种105 kDa的P型ATP酶，它与哺乳动物和真菌的阳离子泵相似。
