The inositol pyrophosphate synthesis pathway in Trypanosoma brucei is linked to polyphosphate synthesis in acidocalcisomes.
The inositol pyrophosphate synthesis pathway in Trypanosoma brucei is linked to polyphosphate synthesis in acidocalcisomes.
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DOI：
10.1111/mmi.13766
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发表时间：
2017-10
影响因子：
3.6
通讯作者：
Docampo R
中科院分区：
文献类型：
--
作者：
Cordeiro CD;Saiardi A;Docampo R
焦磷酸肌醇是一类具有高能焦磷酸键的新型信号分子，具有多种生物学功能。在这里，我们报道了布氏锥虫中与肌醇焦磷酸生物合成途径有关的几种激酶的正确鉴定和特征：肌醇多聚磷酸多激酶(TbIPMK)、肌醇五磷酸2-激酶(TbIP5K)和肌醇六磷酸激酶(TbIP6K)。TbIP5K和TbIP6K不能单独通过序列进行鉴定，它们的活性可以通过重组蛋白的酶活性测定或通过它们与酵母突变株的互补来验证。我们还用聚丙烯酰胺凝胶电泳法和高效液相色谱法分析了布鲁氏毛滴虫提取物中肌醇磷酸盐的存在。有趣的是，在布鲁氏毛滴虫的不同发育阶段，我们都能检测到肌醇磷酸(IP)、4，5-二磷酸(IP2)、1，4，5-三磷酸(IP3)和六磷酸肌醇(IP6)。由于条件性基因敲除下调了TbIPMK的表达，血液中不能产生肌醇焦磷酸盐的形式减少了聚磷酸盐的水平，并改变了酸性钙体。我们的研究将肌醇焦磷酸途径与酸性钙体中多聚磷酸的合成联系起来，可能有助于更好地了解这些生物，并为药物开发提供新的靶点。我们报道了在布鲁氏锥虫中参与肌醇焦磷酸生物合成途径的蛋白激酶的正确鉴定和特征，并鉴定了这些反应的底物和产物。我们通过酵母突变体的互补为它们的活性提供了功能证据，并建立了这一途径与锥虫体内酸性钙体多聚磷酸合成的相关性。
Inositol pyrophosphates are novel signaling molecules possessing high-energy pyrophosphate bonds and involved in a number of biological functions. Here, we report the correct identification and characterization of the kinases involved in the inositol pyrophosphate biosynthetic pathway in Trypanosoma brucei: inositol polyphosphate multikinase (TbIPMK), inositol pentakisphosphate 2-kinase (TbIP5K) and inositol hexakisphosphate kinase (TbIP6K). TbIP5K and TbIP6K were not identifiable by sequence alone and their activities were validated by enzymatic assays with the recombinant proteins or by their complementation of yeast mutants. We also analyzed T. brucei extracts for the presence of inositol phosphates using polyacrylamide gel electrophoresis and high performance liquid chromatography. Interestingly, we could detect inositol phosphate (IP), inositol 4,5-bisphosphate (IP2), inositol 1,4,5-trisphosphate (IP3) and inositol hexakisphosphate (IP6) in T. brucei different stages. Bloodstream forms unable to produce inositol pyrophosphates, due to downregulation of TbIPMK expression by conditional knockout, have reduced levels of polyphosphate and altered acidocalcisomes. Our study links the inositol pyrophosphate pathway to the synthesis of polyphosphate in acidocalcisomes, and may lead to better understanding of these organisms and provide new targets for drug discovery. We report the correct identification and characterization of the kinases involved in the inositol pyrophosphate biosynthetic pathway in Trypanosoma brucei and identified the substrates and products of these reactions. We provide functional evidence of their activity by complementation of yeast mutants, and we establish the relevance of this pathway for acidocalcisome polyphosphate synthesis in trypanosomes.
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从细菌到人类细胞的细胞器保守的酸性蛋白质组学分析。
影响因子：
6.7
作者：
Huang G;Ulrich PN;Storey M;Johnson D;Tischer J;Tovar JA;Moreno SN;Orlando R;Docampo R
通讯作者：
Docampo R
DOI：
10.1073/pnas.0909176106
发表时间：
2009-12-15
影响因子：
11.1
作者：
Azevedo, Cristina;Burton, Adam;Saiardi, Adolfo
通讯作者：
Saiardi, Adolfo
影响因子：
8.6
作者：
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Stuart, Ken
影响因子：
4.8
作者：
Choi, Jae H.;Williams, Jason;Shears, Stephen B.
通讯作者：
Shears, Stephen B.
影响因子：
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作者：
Draskovic, Petra;Saiardi, Adolfo;Podobnik, Marjetka
通讯作者：
Podobnik, Marjetka