Elements of Nucleotide Specificity in the Trypanosoma brucei Mitochondrial RNA Editing Enzyme RET2
Elements of Nucleotide Specificity in the Trypanosoma brucei Mitochondrial RNA Editing Enzyme RET2
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DOI：
10.1021/ci3001327
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发表时间：
2012-05-01
影响因子：
5.6
通讯作者：
Amaro, Rommie E.
中科院分区：
文献类型：
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作者：
Demir, Oezlem;Amaro, Rommie E.
非洲昏睡病的病原体布氏锥虫（Trypanosoma brucei）经历了一个不寻常的线粒体RNA编辑过程，这对其生存至关重要。RNA编辑末端尿苷酰转移酶2。brucei（TbRET 2）是进行这种编辑的编辑体机制的不可或缺的组成部分。TbRET 2是维持昆虫和血液形式的寄生虫的活力所必需的，并且由于其高分辨率的晶体结构，它成为一个有前途的药物靶标。对于尿苷5 '-三磷酸（UTP）催化的唯一要求和TbRET 2的RNA引物偏好都没有很好的理解。使用全原子显式溶剂化分子动力学（MD）模拟，我们研究了TbRET 2的结构和动力学，以及决定因素TbRET 2的独家UTP偏好的UTP绑定的效果。通过我们对各种核苷三磷酸底物（NTPs）的研究，我们发现UTP通过广泛的水介导的氢键网络预组织结合位点，将Glu 424和Arg 144侧链带到RNA引物结合的最佳位置。相反，胞嘧啶5 '-三磷酸（CTP）和腺苷S'-三磷酸（ATP）不能实现这种预组织，因此妨碍了生产性RNA引物结合。此外，我们已经定位的配体结合的“热点”的TbRET 2的基础上的MD构象合奏和计算片段映射。TbRET 2在apo和UTP结合的MD模拟中揭示了不同的结合口袋，这可以作为抑制剂设计的目标。
The causative agent of African sleeping sickness, Trypanosoma brucei, undergoes an unusual mitochondrial RNA editing process that is essential for its survival. RNA editing terminal uridylyl transferase 2 of T. brucei (TbRET2) is an indispensable component of the editosome machinery that performs this editing. TbRET2 is required to maintain the vitality of both the insect and bloodstream forms of the parasite, and with its high-resolution crystal structure, it poses as a promising pharmaceutical target. Neither the exclusive requirement of uridine 5'-triphosphate (UTP) for catalysis, nor the RNA primer preference of TbRET2 is well-understood. Using all-atom explicitly solvated molecular dynamics (MD) simulations, we investigated the effect of UTP binding on TbRET2 structure and dynamics, as well as the determinants governing TbRET2's exclusive UTP preference. Through our investigations of various nucleoside triphosphate substrates (NTPs), we show that UTP preorganizes the binding site through an extensive water-mediated H-bonding network, bringing Glu424 and Arg144 side chains to an optimum position for RNA primer binding. In contrast, cytosine 5'-triphosphate (CTP) and adenosine S'-triphosphate (ATP) cannot achieve this preorganization and thus preclude productive RNA primer binding. Additionally, we have located ligand-binding "hot spots" of TbRET2 based on the MD conformational ensembles and computational fragment mapping. TbRET2 reveals different binding pockets in the apo and UTP-bound MD simulations, which could be targeted for inhibitor design.