METABOLIC SPECIFICITY OF METHYLTHIOCOFORMYCIN FOR MALARIAL ADENOSINE DEAMINASE

甲硫基辅霉素对疟疾腺苷脱氨酶的代谢特异性

• 批准号: 8169275
• 负责人: STEVEN G ALMO
• 金额: $ 0.17万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169275
• 关键词: Adenine Nucleotides; Adenosine; Affinity; Binding; Binding Sites; Catalytic Domain; Coformycin; Complex; Computer Retrieval of Information on Scientific Projects Database; Enzymes; Erythrocytes; Funding; Grant; Human; Hydrogen Bonding; Hydroxyl Radical; Hypoxanthines; Institution; Kinetics; Metabolic; Mutagenesis; Pathway interactions; Pentostatin; Plasmodium; Plasmodium falciparum; Plasmodium gallinaceum; Plasmodium vivax; Purine Nucleotides; Purines; Reporting; Research; Research Personnel; Resources; Source; Specificity; Structure; Substrate Specificity; United States National Institutes of Health; Water; adenosine deaminase; analog; base; purine

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Plasmodium falciparum is a purine auxotroph requiring hypoxanthine as a key metabolic precursor. Erythrocyte adenine nucleotides are the source of the purine precursors, making adenosine deaminase (ADA) a key enzyme in the pathway of hypoxanthine formation in Plasmodium falciparum. Methylthioadenosine (MTA) is one of purine nucleotide substrates for most malarial ADAs, but not for human ADA. The catalytic site specificity of malarial ADAs permits methylthiocoformycin (MT-coformycin) to act as a Plasmodium-specific transition state analogue with low affinity for human ADA [Tyler, P. C., Taylor, E. A., Frohlich, R. G. G., and Schramm, V. L. (2007) J. Am. Chem. Soc. 129, 6872-6879]. The structural basis for MTA and MT-coformycin specificity in malarial ADAs is the subject of speculation. In this study, we report the crystal structure of ADA from Plasmodium vivax (PvADA) in a complex with MT-coformycin that reveals an unprecedented binding geometry for 5'-methylthioribosyl groups in the malarial ADAs. Compared to malarial ADA complexes with adenosine or deoxycoformycin, 5'-methylthioribosyl groups are rotated 130 degrees . A hydrogen bonding network between Asp172 and the 3'-hydroxyl of MT-coformycin is essential for recognition of the 5'-methylthioribosyl group. Water occupies the 5'-hydroxyl binding site when MT-coformycin is bound. Mutagenesis of Asp172 destroys the substrate specificity for MTA and MT-coformycin. Kinetic, mutagenic, and structural analyses of PvADA and kinetic analysis of five other Plasmodium ADAs establish the unique structural basis for its specificity for MTA and MT-coformycin. Plasmodium gallinaceum ADA does not use MTA as a substrate, is not inhibited by MT-coformycin, and is missing Asp172.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 恶性疟原虫是一种嘌呤营养缺陷型，需要次黄嘌呤作为关键的代谢前体。红细胞腺嘌呤核苷酸是嘌呤前体的来源，使得腺苷脱氨酶 (ADA) 成为恶性疟原虫次黄嘌呤形成途径中的关键酶。甲硫腺苷 (MTA) 是大多数疟疾 ADA 的嘌呤核苷酸底物之一，但不是人类 ADA。疟疾 ADA 的催化位点特异性使得甲硫基辅福霉素 (MT-coformycin) 可以作为疟原虫特异性过渡态类似物，对人 ADA 具有低亲和力 [Tyler, P. C.、Taylor, E. A.、Frohlich, R. G. G. 和 Schramm, V. L. (2007) J. Am.化学。苏克。 129、6872-6879]。疟疾 ADA 中 MTA 和 MT-辅福霉素特异性的结构基础是推测的主题。在这项研究中，我们报道了来自间日疟原虫 (PvADA) 的 ADA 与 MT-辅福霉素复合物的晶体结构，揭示了疟疾 ADA 中 5'-甲基硫核糖基的前所未有的结合几何结构。与疟疾 ADA 与腺苷或脱氧考福霉素的复合物相比，5'-甲基硫核糖基旋转了 130 度。 Asp172 和 MT-coformycin 3'-羟基之间的氢键网络对于识别 5'-甲基硫核糖基至关重要。当 MT-辅福霉素结合时，水占据 5'-羟基结合位点。 Asp172 的诱变破坏了 MTA 和 MT-coformycin 的底物特异性。 PvADA 的动力学、诱变和结构分析以及其他五种疟原虫 ADA 的动力学分析为其对 MTA 和 MT-coformycin 的特异性奠定了独特的结构基础。鸡疟原虫 ADA 不使用 MTA 作为底物，不受 MT-辅福霉素抑制，并且缺少 Asp172。