Two classes of plant-like vacuolar-type H+-pyrophosphatases in malaria parasites
Two classes of plant-like vacuolar-type H+-pyrophosphatases in malaria parasites
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DOI：
10.1016/s0166-6851(01)00251-1
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发表时间：
2001-05-01
影响因子：
1.5
通讯作者：
Vaidya, AB
中科院分区：
文献类型：
--
作者：
McIntosh, MT;Drozdowicz, YM;Vaidya, AB
在植物中，胞质无机焦磷酸盐（PP）由节能型液泡型H+-焦磷酸酶（V-PPases）水解，该酶利用PP水解的自由能建立跨膜H+梯度。本文描述了疟原虫恶性疟原虫PfVP1和PfVP2两个基因的鉴定和克隆。据推测，PfVP1和PfVP2分别编码I型（依赖K+）和II型（不依赖K+） v - ppase，在植物中，它们之间的序列差异比它们与I型和II型的序列差异更大。与PfVP2 mRNA相比，PfVP1 mRNA的稳态水平在感染的整个红细胞期都很高。用通用V-PP对滋养体膜进行Western分析ise抗体（PAB（HK）和PAB(TK)）显示出相当数量的Mr 67000多肽，其相关的氨基二膦酸盐（AMDP）抑制PPase活性被K+显著刺激。感染红细胞的免疫荧光显微镜显示PfVP抗原与寄生虫的质膜和点状细胞内包涵体相关。瞬时转染PfVP1 - gfp融合进一步支持PfVP1在寄生虫质膜上的定位。基于这些发现和AMDP的生长抑制作用，我们认为恶性疟原虫具有I型和II型v - ppase，其中在能量限制条件下，前者最有可能促进H+在寄生虫质膜上的梯度建立。(C) 2001 Elsevier Science B.V.版权所有
In plants, cytosolic inorganic pyrophosphate (PP,) is hydrolyzed by energy-conserving vacuolar-type H+-pyrophosphatases (V-PPases) that harness the free energy of PP, hydrolysis to establish transmembrane H+ gradients. Here we describe the identification and cloning of two genes, PfVP1 and PfVP2, from the malaria parasite Plasmodium falciparum. Inferred to encode type I (K+-dependent) and type II (K+-independent) V-PPases, respectively, PfVP1 and PfVP2 appeared more sequence divergent from each other than from their type I and type II counterparts in plants. The steady state levels of PfVP1 mRNA were high in comparison to PfVP2 mRNA throughout the erythrocytic phases of infection. Western analyses of trophozoite membranes using generic V-PP;ise antibodies (PAB(HK) and PAB(TK)) demonstrated appreciable amounts of a Mr 67000 polypeptide whose associated aminomethylenediphosphonate- (AMDP) inhibitable PPase activity was markedly stimulated by K+. Immunofluorescence microscopy of infected erythrocytes revealed PfVP antigen associated with both the parasite plasma membrane and punctate intracellular inclusions. Transient transfection of a pfVP1-GFP fusion further supported the localization of PfVP1 to the parasite plasma membrane. Based on these findings and the growth-retarding effects of AMDP, P. falciparum is concluded to possess both type I and type II V-PPases of which thr former has the greatest potential for contributing to the establishment of H+ gradients across the parasite plasma membrane under conditions of energy limitation. (C) 2001 Elsevier Science B.V. All rights reserved.