Inhibitors of Trypanothione Reductase

描述（由申请人提供）：本提案重点关注锥虫硫酮还原酶（TR）潜在抑制剂的设计、合成和评价。TR是一种NADPH依赖性黄素酶，可催化抗氧化剂锥虫硫酮（N1，N8-双（谷胱甘肽）亚精胺）从其二硫化物还原为二硫醇形式。这种酶存在于锥虫寄生虫、锥虫属和利什曼原虫中，它们是人类和家畜的许多疾病的病原体。实例包括非洲昏睡病（布氏锥虫）、恰加斯病（T. cruzi），以及许多利什曼原虫感染，如东方疮（热带利什曼原虫）和黑热病（L. Donovani）。这些疾病对健康和经济的影响是巨大的;然而，缺乏真正有效的治疗方法。自20世纪80年代中期发现以来，TR已成为抗寄生虫药物开发的主要目标。许多注意力集中在可逆TR抑制剂的鉴定上。虽然这项研究将增加一些建议的竞争性TR抑制剂的知识体系，其主要重点是确定基于机制的不可逆抑制剂，一类TR抑制剂，在文献中很少受到关注。在所提出的不可逆抑制剂的设计中所采取的方法是制备将亲电子部分代替底物的二硫化物基团的锥虫硫酮类似物。预计亲电抑制剂将与酶活性位点中的亲核半胱氨酸残基形成共价键，该残基已知是TR催化机制中的关键残基。实验方法将是有机化学，特别是核磁共振（NMR）光谱和电喷雾电离（ESI）质谱将在制备和分析所提出的化合物中发挥重要作用。将使用动力学和质谱方法评价潜在抑制剂的抑制活性。特别是，通过HPLC/ESI离子阱MS/MS对抑制蛋白的分析在验证所提出的不可逆抑制剂的共价性质方面非常重要。卡尔顿学院本科化学专业将开展这项研究。首席研究员在制作成功的本科生研究经验方面有着丰富的经验。

DESCRIPTION (provided by applicant): This proposal focuses on the design, synthesis, and evaluation of potential inhibitors of trypanothione reductase (TR). TR is an NADPH-dependent flavoenzyme that catalyzes the reduction of the antioxidant trypanothione (N1, N8-bis (glutathionyl) spermidine) from its disulfide to its dithiol form. The enzyme is found in the trypanosomatid parasites, Trypanosoma and Leishmania, causative agents of a host of diseases of both humans and domestic animals. Examples include African sleeping sickness (Trypanosoma brucei), Chagas' disease (T. cruzi), and numerous leishmanial infections, such as oriental sore (Leishmania tropica) and kalazar (L. donovani). The health and economic effects of these diseases are enormous; however, truly effective treatments for them are lacking. Since its discovery in the mid-1980s, TR has emerged as a prime target for the development of antiparasitic drugs. Much attention has been focused on the identification of reversible TR inhibitors. While this study will add to that body of knowledge with a number of proposed competitive TR inhibitors, its primary focus is to identify mechanism-based irreversible inhibitors, a class of TR inhibitor that has received scant attention in the literature. The approach taken in the design of the proposed irreversible inhibitors is to prepare analogues of trypanothione that incorporate electrophilic moieties in place of the substrate's disulfide group. It is expected that the electrophilic inhibitors will form a covalent bond with a nucleophilic cysteine residue in the enzyme active site, which is known to be a key residue in the catalytic mechanism of TR. The experimental methods will be those of organic chemistry, in particular, nuclear magnetic resonance (NMR) spectroscopy and electro spray ionization (ESI) mass spectrometry will play a large role in the preparation and analysis of the proposed compounds. The inhibitory activity of potential inhibitors will be evaluated using both kinetic and mass spectrometric approaches. In particular, analysis of inhibited protein via HPLC/ESI ion trap MS/MS will be important in verifying the covalent nature of the proposed irreversible inhibitors. Carleton College undergraduate chemistry majors will carry out the research. The principal investigator has had significant experience in crafting successful undergraduate research experiences.