Structure and mechanism of tRNA splicing enzymes Trl1 and Tpt1: therapeutic targets for fungal infection.

tRNA 剪接酶 Trl1 和 Tpt1 的结构和机制：真菌感染的治疗靶点。

• 批准号: 394320208
• 负责人: Dr. Ankan Banerjee
• 金额: --
• 依托单位: Molecular Biology Program
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/394320208?language=en
• 关键词: Structure; mechanism; tRNA; splicing; enzymes

Invasive fungal infections, especially Aspergillosis and Candidiasis, are a major cause of morbidity and mortality in cancer patients with prolonged neutropenia following chemotherapy or hematopoietic stem cell transplantation. The development of more effective (and less toxic) drugs for the treatment of fungal infections in cancer settings hinges on defining new targets for antifungal drug discovery and implementing strategies to identify inhibitors. Although the tRNA splicing enzymes are promising anti-fungal drug targets, a prime bottleneck is the lack of atomic structures for the unique fungal tRNA ligase (Trl1) and 2´-phosphotransferase (Tpt1). To that end, I intend to pursue structural, biochemical and genetic characterization of the Trl1 ligase domain and Tpt1 from the fungal pathogens Aspergillus fumigatus and Candida albicans. The premise of the project is that clarifying the structures and mechanisms of the Aspergillus and Candida tRNA splicing enzymes will drive progress toward drug discovery. The specific aims for the 2-year project proposal are: 1. To crystallize and determine the atomic structures of the Trl1- ligase domain of fungal pathogens, in complexes with substrates, cofactors, and reaction intermediates. 2. To determine crystal structures of Tpt1 in complexes with substrates (NAD+ and a synthetic RNA with a 3´-5´-phosphodiester, 2´-phosphate splice junction) and to trap Tpt1 in a complex with the RNA 2´- phospho-ADP-ribosylated reaction intermediate.This will fill a major knowledge gap in tRNA metabolism by providing insights to involved active site architectures and catalytic mechanisms. The structure-function information will promote the discovery of antifungals.

侵袭性真菌感染，特别是曲霉病和念珠菌病，是化疗或造血干细胞移植后长期中性粒细胞减少的癌症患者发病率和死亡率的主要原因。开发更有效（且毒性更低）的药物用于治疗癌症环境中的真菌感染，取决于确定抗真菌药物发现的新目标和实施识别抑制剂的策略。虽然tRNA剪接酶是有希望的抗真菌药物靶点，但主要瓶颈是缺乏独特的真菌tRNA连接酶（Trl 1）和2 ′-磷酸转移酶（Tpt 1）的原子结构。为此，我打算追求的Trl 1连接酶结构域和Tpt 1的真菌病原体烟曲霉和白色念珠菌的结构，生化和遗传特性。该项目的前提是澄清曲霉和念珠菌tRNA剪接酶的结构和机制将推动药物发现的进展。2年项目建议书的具体目标是：1。 结晶和确定真菌病原体的Trl 1-连接酶结构域的原子结构，与底物，辅因子和反应中间体的复合物。2. 确定Tpt 1与底物（NAD+和具有3 ′-5 ′-磷酸二酯，2 ′-磷酸剪接点的合成RNA）复合物的晶体结构，并将Tpt 1捕获在与RNA 2 ′-磷酸-ADP-核糖基化反应中间体的复合物中。这将通过提供对所涉及的活性位点结构和催化机制的见解来填补tRNA代谢的主要知识空白。这些结构-功能信息将促进抗真菌药物的发现。

项目成果

Atomic structures of the RNA end-healing 5′-OH kinase and 2′,3′-cyclic phosphodiesterase domains of fungal tRNA ligase: conformational switches in the kinase upon binding of the GTP phosphate donor

• DOI: 10.1093/nar/gkz1049
• 发表时间: 2019-12-16
• 期刊: NUCLEIC ACIDS RESEARCH
• 影响因子: 14.9
• 作者: Banerjee, Ankan;Goldgur, Yehuda;Shuman, Stewart
• 通讯作者: Shuman, Stewart