BIOENERGETICS AND PROTON PUMPS IN MALARIA PARASITES

描述(由申请人提供)：跨质膜运输是所有细胞生理学的重要特征。质膜上的许多守门人是由跨膜的离子和电梯度辅助的。细胞消耗巨大的能量--高达细胞内ATP的50%--来维持这种电化学梯度。在疟疾寄生虫中，用电化学梯度给质膜充电以进行这种运输的能量经济性几乎仍然未知。事实上，疟疾寄生虫的红细胞期主要通过底物水平的磷酸化来获得ATP，每个葡萄糖分子只能勉强维持两个ATP分子，这肯定会对寄生虫的能量利用造成很大的限制。该项目旨在探索疟疾寄生虫的替代和/或附属能源。最近的证据表明，疟原虫包含两个成员(PfVP1和PfVP2)，它们是类植物节能的、膜相关的H~*泵焦磷酸酶。植物的液泡焦磷酸酶(V-PPase)结合了无机焦磷酸(PPI)的磷酸酸化键(PPI)水解产生的能量，从而将Hv泵送到液泡膜上。在疟疾寄生虫中，初步数据表明该酶位于寄生虫的质膜内。这一位置表明，Hv跨寄生虫质膜的转运可能通过PPase的PPI水解和V型ATPase的ATP水解而被激活。由于动物细胞不具有V-PPase的同源物，这些酶在疟疾寄生虫中的存在为设计选择性毒性抑制剂提供了候选。该项目将对PfVP1和PfVP2的生物化学和细胞生物学进行基础研究。将采用基因破坏方法来评估这些分子对寄生虫生理学的贡献。探讨疟疾寄生虫V-ATPase在高能需求下，利用V-PPase产生的质子动力反向合成ATP的可能性。此外，将研究观察到的疟疾寄生虫FoF1-ATP合成酶的一个不寻常的亚基配置，以评估这种通常是线粒体质子泵复合体对寄生虫生理的贡献。该项目的结果可能需要对我们对疟疾寄生虫生物能量学的看法进行重大修订。该项目中可能发现的疟疾寄生虫中质子动态平衡和生物能量学的独特特征可能成为设计新的疟疾控制方法的基础。

DESCRIPTION (provided by the applicant): Transport across the plasma membrane is a critical feature of all cellular physiology. Many gatekeepers at the plasma membrane are assisted by ionic and electric gradients across the membrane. Cells expend enormous energy-- up to 50% of intracellular ATP-- for maintenance of such electrochemical gradients. Energy economics of charging the plasma membrane with an electrochemical gradient for such transport remains virtually unknown in malaria parasites. The fact that erythrocytic stages of malaria parasites derive their ATP mainly through substrate-level phosphorylation, eking out a mere two ATP molecules per glucose molecule, must place significant constraints on parasite energy utilization. This project seeks to explore an alternate and/or adjunct energy source for malaria parasites. Recent evidence shows that Plasmodium species contains two members (PfVP1 and PfVP2) of the plant-like energyconserving, membrane-associated H*-pumping pyrophosphatases. The vacuolar pyrophosphatases (V-PPases) of plants couple the energy generated by hydrolysis of the phosphoanhydride bond of inorganic pyrophosphate (PPi) to pump Hv across the vacuolar membrane. In malaria parasites, preliminary data suggest that the enzyme is located within the parasite plasma membrane. This location would suggest that Hv translocation across the parasite plasma membrane could be energized through PPi hydrolysis by the PPases in concert with ATP hydrolysis by the V-type ATPase. Because animal cells do not possess homologues of V-PPases, the presence of these enzymes in malaria parasites offers candidates for devising selectively toxic inhibitors. This project will undertake basic investigations on the biochemistry and cell biology of PfVP1 and PfVP2. Gene disruption approaches will be undertaken to assess contributions made by these molecules to the parasite physiology. The possibility that V-ATPase of malaria parasites may function work in reverse to synthesize ATP by using the proton motive force generated by the V-PPases under high-energy demand will be explored. Furthermore, an unusual subunit configuration observed for the FoF1-ATP synthase of malaria parasites will be investigated to assess the contribution of this usually mitochondrial proton pumping complex to parasite physiology. Results from this project have a potential to require a major revision of our view of malaria parasite bioenergetics. Unique features of proton homeostasis and bioenergetics in malaria parasites likely to be uncovered in this project could form the basis for devising novel approaches to malaria control.