Ligand-bound structures provide atomic snapshots for the catalytic mechanism of D-amino acid deacylase.
Ligand-bound structures provide atomic snapshots for the catalytic mechanism of D-amino acid deacylase.
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DOI：
10.1074/jbc.m109.038562
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发表时间：
2010-02-19
期刊：
影响因子：
--
通讯作者：
Sharma A
中科院分区：
文献类型：
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作者：
Bhatt TK;Yogavel M;Wydau S;Berwal R;Sharma A
D-酪氨酰-tRNatyr脱酰酶(DTD)是一种从错误充电的tRNA中去除d-氨基酸的编辑酶。我们在这里描述了对疟疾寄生虫恶性疟原虫DTD的深入分析。我们的数据为DTD与腺苷和d-氨基酸的络合物提供了结构洞察力。结合的腺苷位于DTD催化位点的近端，它代表带电tRNA的真实末端腺苷。DTD结合的d-氨基酸聚集在整个活性位点口袋中的三个不同的亚位点上。这些亚位称为过渡亚位、活性亚位和退出亚位，允许对接、重定向、手性选择、催化和游离d-氨基酸从DTD中退出。我们的研究揭示了DTD识别d-氨基酸的不同模式，表明了一种固有的可塑性，可以容纳所有d-氨基酸。对天然的、ADP结合的和d-氨基酸络合的DTD结构的深入分析提供了该酶家族识别配体和随后催化的第一个原子快照。我们已经绘制了脱酰化反应的位置，并标记了所有底物和产物进入和离开的可能路线。我们还进行了基于结构的抑制剂发现，并使用生长抑制试验测试了先导化合物对疟疾寄生虫恶性疟原虫的抑制作用。我们的研究为DTD酶的催化机制提供了全面的结构基础，并对抑制恶性疟原虫中的DTD酶作为抑制寄生虫的一种途径具有重要意义。
d-tyrosyl-tRNATyr deacylase (DTD) is an editing enzyme that removes d-amino acids from mischarged tRNAs. We describe an in-depth analysis of the malaria parasite Plasmodium falciparum DTD here. Our data provide structural insights into DTD complexes with adenosine and d-amino acids. Bound adenosine is proximal to the DTD catalysis site, and it represents the authentic terminal adenosine of charged tRNA. DTD-bound d-amino acids cluster at three different subsites within the overall active site pocket. These subsites, called transition, active, and exit subsites allow docking, re-orientation, chiral selection, catalysis, and exit of the free d-amino acid from DTD. Our studies reveal variable modes of d-amino acid recognition by DTDs, suggesting an inherent plasticity that can accommodate all d- amino acids. An in-depth analysis of native, ADP-bound, and d- amino acid-complexed DTD structures provide the first atomic snapshots of ligand recognition and subsequent catalysis by this enzyme family. We have mapped sites for the deacylation reaction and mark possible routes for entry and egress of all substrates and products. We have also performed structure-based inhibitor discovery and tested lead compounds against the malaria parasite P. falciparum using growth inhibition assays. Our studies provide a comprehensive structural basis for the catalytic mechanism of DTD enzymes and have implications for inhibition of this enzyme in P. falciparum as a route to inhibiting the parasite.