Institute of Cell Biology, University of Bern, Baltzerstrasse 4, CH-3012 Bern, Switzerland.
BMC Microbiol. 2011 Jan 6;11:4. doi: 10.1186/1471-2180-11-4.
Exopolyphosphatases and pyrophosphatases play important but still incompletely understood roles in energy metabolism, and also in other aspects of cell biology such as osmoregulation or signal transduction. Earlier work has suggested that a human exopolyphosphatase, Prune, might exhibit cyclic nucleotide phosphodiesterase activity.
The kinetoplastida, a large order of unicellular eukaryotes that contains many important pathogens such as Trypanosoma brucei (human sleeping sickness), Trypanosoma cruzi (Chagas disease) or Leishmania ssp (several clinically dinstinct leishmaniases) all contain several exo- and pyrophosphatases. The current study provides a systematic classification of these enzymes, which now allows to situate the information that is already available on some of these enzymes. It then analyses the exopolyphosphatase TbrPPX1 of T. brucei in detail, using RNA interference and genetic knockouts in an attempt to define its function, and immunofluorescence microscopy to study its subcellular localization.TbrPPX1 is an exopolyphosphatase that does hydrolyze pentasodium triphosphate, but not organic triphosphates such as ATP, pyrophosphate or long-chain polyphosphates. Finally, the study investigates the potential cyclic nucleotide phosphodiesterase activity of TbrPPX1.
All kinetoplastid genomes that are currently available contain genes for an exopolyphosphatase and two classes of pyrophosphatases, one associated with the acidocalcisomes and one cytoplasmic. TbrPPX1 represents the T. brucei exopolyphosphatase. It is located throughout the cytoplasm, and its genetic ablation does not produce a dramatic phenotype. Importantly, TbrPPX1 does not exhibit any cyclic nucleotide specific phosphodiesterase activity, which definitively eliminates it as an additional player in cAMP signalling of the kinetoplastida.
外切多聚磷酸酶和焦磷酸酶在能量代谢中发挥着重要但仍不完全了解的作用，在细胞生物学的其他方面也发挥着重要作用，如渗透调节或信号转导。早期的工作表明，人类外切多聚磷酸酶 Prune 可能具有环核苷酸磷酸二酯酶活性。
动基体门，一个由单细胞真核生物组成的大目，其中包含许多重要的病原体，如布氏锥虫（人类昏睡病）、克氏锥虫（恰加斯病）或利什曼原虫（几种不同的利什曼病），都含有几种外切和焦磷酸酶。本研究对这些酶进行了系统分类，现在可以将已经获得的关于其中一些酶的信息定位。然后，它详细分析了布氏锥虫的外切多聚磷酸酶 TbrPPX1，使用 RNA 干扰和遗传敲除试图定义其功能，并通过免疫荧光显微镜研究其亚细胞定位。TbrPPX1 是一种外切多聚磷酸酶，可水解五钠三磷酸，但不能水解有机三磷酸，如 ATP、焦磷酸或长链多磷酸。最后，该研究调查了 TbrPPX1 潜在的环核苷酸磷酸二酯酶活性。
目前所有可用的动基体门基因组都包含一个外切多聚磷酸酶和两类焦磷酸酶的基因，一类与酸钙体有关，另一类与细胞质有关。TbrPPX1 代表布氏锥虫的外切多聚磷酸酶。它位于细胞质的各个部位，其基因缺失不会产生明显的表型。重要的是，TbrPPX1 没有表现出任何环核苷酸特异性磷酸二酯酶活性，这明确排除了它作为动基体门 cAMP 信号的另一个参与者。
