Metalloenzyme structure, function and assembly

金属酶的结构、功能和组装

• 批准号: 10621553
• 负责人: CATHERINE L DRENNAN
• 金额: $ 41.03万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10621553
• 关键词: Amaze; Anabolism; Antibiotic Resistance; Antibiotics; Antineoplastic Agents; Antiviral Agents; Architecture; Biochemistry; Biological Models; Biotechnology; Carbon; Carbon Dioxide; Chemicals; Chemistry; Complex; Cryoelectron Microscopy; Crystallography; DNA biosynthesis; Data; Deoxyribonucleotides; Electron Transport; Enzymes; Gases; Hand; Health; Heart; Human; Ions; Life; Medical; Metalloproteins; Metals; Methane; Methods; Molecular; Molecular Conformation; Nature; Organometallic Chemistry; Production; Protein Engineering; Proteins; Reaction; Resolution; Ribonucleotide Reductase; Ribonucleotides; Scaffolding Protein; Structure; System; Toxic Environmental Substances; X-Ray Crystallography; anti-cancer; carbon compound; climate change; combat; design; frontier; greenhouse gases; improved; insight; metalloenzyme; microbial; novel; remediation; sample fixation; scaffold

Abstract The combination of metal ions with proteins offers unique chemical reactivities that are at the heart of many of Nature’s most important and amazing chemical transformations. For example, metalloenzymes catalyze the reduction of ribonucleotides to deoxyribonucleotides, a rate-limiting step in DNA biosynthesis. They biosynthesize anticancer and antiviral compounds that have unusual scaffolds and catalyze the carbon-carbon bond forming reactions that afford life on CO2 and H2 gases. Our lab employs structural methods to interrogate how metalloenzymes are able to perform this incredible chemistry. We seek to understand how the architecture of metalloenzymes allows for radical species to be controlled – i.e. turned off, turned on and harnessed – to enable the reaction at hand. We also strive to understand how proteins are designed to enable long-range electron transfer without protein damage or radical loss. In this proposal, we describe structural studies of our metalloenzyme model systems, including class Ia (diiron-dependent) and class III (glycyl radical- dependent) ribonucleotide reductases that allow us to interrogate the molecular basis of radical-based chemistry. We also describe efforts to understand how protein scaffolds facilitate organometallic chemistry, especially in regard to microbial carbon dioxide fixation and methane production. These studies leverage both our expertise in working with O2-sensitive metalloenzymes and in cryogenic-electron microscopy (cryo-EM). Although we will continue to employ X-ray crystallography, cryo-EM is proving to be a game-changer for many of our metalloenzyme systems. In particular, the resolution revolution of cryo-EM provides us with the means to obtain long-awaited structures of both large (2000 kDa) and transient metalloprotein complexes and to determine structures of metalloenzymes in functionally-essential conformational states that were previously unattainable by crystallography. The results of our structural studies will enable structure-based design of novel antibiotics targeting, for example, microbial ribonucleotide reductases. These structural data will also guide efforts to exploit radical enzymes for the production of medically important compounds with unusual scaffolds. These data will additionally facilitate the application of metalloenzymes or their bioinspired inorganic counterparts in the production of high-value carbon compounds from the greenhouse gas CO2 and, ideally, improve our understanding of some of the more enigmatic aspects of metalloprotein biochemistry.

摘要 金属离子与蛋白质的结合提供了独特的化学反应性，这是许多化学反应的核心。 自然界最重要和最令人惊奇的化学变化。例如，金属酶催化 核糖核苷酸还原为脱氧核糖核苷酸，这是DNA生物合成中的限速步骤。他们 生物合成抗癌和抗病毒化合物，具有不寻常的支架和催化碳-碳 键形成反应，提供了对CO2和H2气体的生命。我们的实验室采用结构化方法来审问 金属酶是如何进行这种不可思议的化学反应的。我们试图了解 金属酶的结构允许自由基种类被控制-即关闭、打开和 利用-使手头的反应。我们还努力了解蛋白质是如何设计的， 长距离电子转移而没有蛋白质损伤或自由基损失。在本建议中，我们描述了结构 我们的金属酶模型系统的研究，包括Ia类（二铁依赖性）和III类（甘氨酰自由基- 依赖的）核糖核苷酸还原酶，使我们能够询问基于自由基的分子基础 化学.我们还描述了理解蛋白质支架如何促进有机金属化学的努力， 特别是在微生物二氧化碳固定和甲烷生产方面。这些研究利用了 我们在O2敏感金属酶和低温电子显微镜（cryo-EM）方面的专业知识。 虽然我们将继续采用X射线晶体学，但冷冻EM被证明是许多人的游戏规则改变者 我们的金属酶系统。特别是，cryo-EM的分辨率革命为我们提供了 获得期待已久的大（2000 kDa）和瞬时金属蛋白复合物的结构， 确定金属酶的结构在功能上必要的构象状态，以前 晶体学无法达到的。我们的结构研究结果将使基于结构的设计， 靶向例如微生物核糖核苷酸还原酶的新抗生素。这些结构数据也将 指导开发自由基酶的努力，以生产具有特殊重要医学意义的化合物。 脚手架这些数据将另外促进金属酶或其生物启发的无机酶的应用。 从温室气体CO2中生产高价值碳化合物的同行，理想情况下， 提高我们对金属蛋白生物化学中一些更神秘的方面的理解。

项目成果

Starting a new chapter on class Ia ribonucleotide reductases.

• DOI: 10.1016/j.sbi.2022.102489
• 发表时间: 2022-10
• 期刊: Current opinion in structural biology
• 影响因子: 6.8
• 作者: T. Levitz;C. Drennan

Structural and spectroscopic analyses of the sporulation killing factor biosynthetic enzyme SkfB, a bacterial AdoMet radical sactisynthase.

• DOI: 10.1074/jbc.ra118.005369
• 发表时间: 2018-11-09
• 期刊: The Journal of biological chemistry
• 作者: Grell TAJ;Kincannon WM;Bruender NA;Blaesi EJ;Krebs C;Bandarian V;Drennan CL
• 通讯作者: Drennan CL

Development of an in vitro method for activation of X-succinate synthases for fumarate hydroalkylation.

• DOI: 10.1016/j.isci.2023.106902
• 发表时间: 2023-06-16
• 期刊: ISCIENCE
• 影响因子: 5.8
• 作者: Andorfer, Mary C.;King-Roberts, Devin T.;Imrich, Christa N.;Brotheridge, Balyn G.;Drennan, Catherine L.
• 通讯作者: Drennan, Catherine L.

The Atypical Cobalamin-Dependent S-Adenosyl-l-Methionine Nonradical Methylase TsrM and Its Radical Counterparts.

• DOI: 10.1021/jacs.1c12064
• 发表时间: 2022-04-06
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Ulrich EC;Drennan CL
• 通讯作者: Drennan CL

Structural and biochemical investigations of a HEAT-repeat protein involved in the cytosolic iron-sulfur cluster assembly pathway.

• DOI: 10.1038/s42003-023-05579-3
• 发表时间: 2023-12-18
• 期刊: COMMUNICATIONS BIOLOGY
• 影响因子: 5.9
• 作者: Vasquez, Sheena;Marquez, Melissa D.;Brignole, Edward J.;Vo, Amanda;Kong, Sunnie;Park, Christopher;Perlstein, Deborah L.;Drennan, Catherine L.
• 通讯作者: Drennan, Catherine L.