LEISHMANIA PROTON PUMP: GENES & PROTEINS

多诺瓦尼乳杆菌的主动转运依赖于质子梯度 穿过通过动作建立起来的质膜 一种能释放质子的ATPase。这个质子一定也起到了 在酸性环境中寄生虫的生存起着至关重要的作用 脊椎动物宿主的吞噬溶酶体的环境。我们的长期计划 目的是了解肌动蛋白的结构、功能和调节 这种酶，并通过这种理解提供了途径 对这种重要病原体的控制。这项建议的目的是 就是应用分子克隆技术来生产分子 用来分析大分子的探针，这些大分子参与了 质子泵系统。我们在第一阶段取得了成功 这项努力已经克隆了两个排列有序的基因的一部分 它们在结构上能够编码质膜阳离子 打气筒。我们将克隆这对串联的多诺瓦尼乳杆菌基因 他们的全部。我们将确定这些基因的DNA序列， 并预测其蛋白质产物的氨基酸序列。 这些序列信息将被用来生产核酸和 用于分析两种生命中表达的抗体探针 寄生虫的周期阶段。我们将会比较 这些基因产物对质子泵的活性，并检测 推测的编码蛋白的酶活性 质子泵基因。

Active transport in L. donovani is dependent on a proton gradient across the plasma membrane that is established through the action of an ATPase that extrudes protons. This proton must also play a vital role in the survival of the parasite within the acidic environment of the vertebrate host's phagolysosomes. Our long term goal is to understand the structure, function and regulation of this enzyme and, through this understanding to provide avenues to the control of this important pathogen. The aim of this proposal is to apply molecular cloning technology to produce molecular probes with which to analyze the macromolecules that mediate the proton pumping system. We have succeeded in the first phase of this endeavor and have cloned parts of two tandemly arrayed genes that are structurally capable of encoding plasma membrane cation pumps. We will clone this tandem pair of L. donovani genes in their entirety. We will determine the DNA sequences of the genes, and predict the amino acid sequences of their protein products. This sequence information will be used produce nucleic acid and antibody probes for the analysis of expression in the two life cycle stages of the parasite. We will compare the abundance of these gene products to proton pumping activity, and examine the enzymatic activities of the proteins encoded by the presumptive proton pump genes.