Iron-dependent RNA-binding activity of Mycobacterium tuberculosis aconitase
Iron-dependent RNA-binding activity of Mycobacterium tuberculosis aconitase
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DOI：
10.1128/jb.00026-07
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发表时间：
2007-06-01
影响因子：
3.2
通讯作者：
Hasnain, Seyed E.
中科院分区：
文献类型：
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作者：
Banerjee, Sharmistha;Nandyala, Ashok Kumar;Hasnain, Seyed E.
细胞铁水平由结核分枝杆菌的铁调节和传感器蛋白密切监测，以在巨噬细胞内存活。在真核生物中系统研究并在少数原核生物中报道的一类蛋白质是铁响应蛋白（IRP）。这些IRP与mRNA非翻译区（UTR）的铁响应元件（IRE）结合，负责转录后调节参与铁稳态的蛋白质表达。M.结核乌头酸酶（Acn），一种三羧酸（TCA）循环酶，显示存在IRP类蛋白的保守残基。我们证明了M.结核Acn是双功能的。它是一种单体蛋白质，在7至10的宽pH范围内（最佳pH 8），在将异柠檬酸盐转化为顺乌头酸盐方面具有酶活性。从凝胶阻滞试验中可以明显看出，M.结核Acn也通过结合已知的哺乳动物IRE样序列和预测的存在于M的硫氧还蛋白（trxC）的3'UTR和铁依赖性阻遏物和激活物（ideR）的5'UTR的IRE样序列而表现出类似IRP的行为。结核M.当用Fe 2+再活化时，结核杆菌Acn作为TCA循环酶发挥功能，但在通过特异性铁螯合剂消耗铁时，其表现得像IRP，在体外结合所选择的IRE。由于铁是Acn活性所必需的，并抑制Acn的RNA结合活性，因此M.结核病Acn是相互排斥的。我们的研究结果证明了M的双功能性质。结核性痤疮，指出其可能在铁稳态的作用。
Cellular iron levels are closely monitored by iron regulatory and sensor proteins of Mycobacterium tuberculosis for survival inside macrophages. One such class of proteins systematically studied in eukaryotes and reported in a few prokaryotes are the iron-responsive proteins (IRPs). These IRPs bind to iron-responsive elements (IREs) present at untranslated regions (UTRs) of mRNAs and are responsible for posttranscriptional regulation of the expression of proteins involved in iron homeostasis. Amino acid sequence analysis of M. tuberculosis aconitase (Acn), a tricarboxylic acid (TCA) cycle enzyme, showed the presence of the conserved residues of the IRP class of proteins. We demonstrate that M. tuberculosis Acn is bifunctional. It is a monomeric protein that is enzymatically active in converting isocitrate to cis-aconitate at a broad pH range of 7 to 10 (optimum, pH 8). As evident from gel retardation assays, M. tuberculosis Acn also behaves like an IRP by binding to known mammalian IRE-like sequences and to predicted IRE-like sequences present at the 3' UTR of thioredoxin (trxC) and the 5' UTR of the iron-dependent repressor and activator (ideR) of M. tuberculosis. M. tuberculosis Acn when reactivated with Fe2+ functions as a TCA cycle enzyme, but upon iron depletion by a specific iron chelator, it behaves like an IRP, binding to the selected IREs in vitro. Since iron is required for the Acn activity and inhibits the RNA-binding activity of Acn, the two activities of M. tuberculosis Acn are mutually exclusive. Our results demonstrate the bifunctional nature of M. tuberculosis Acn, pointing to its likely role in iron homeostasis.