Unraveling the principles of catalytic diversity in the carotenoid oxygenase superfamily

揭示类胡萝卜素加氧酶超家族催化多样性的原理

• 批准号: 2107713
• 负责人: Philip Kiser
• 金额: $ 52.8万
• 依托单位: University of California-Irvine
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2107713&HistoricalAwards=false
• 关键词: Unraveling; principles; catalytic; diversity; carotenoid

With the support of the Chemistry of Life Processes Program in the Division of Chemistry, Dr. Philip Kiser from the University of California, Irvine is studying a family of enzymes known as carotenoid cleavage dioxygenases (CCDs). These enzymes have a highly conserved 4-His iron center and exhibit a remarkably diverse set of catalytic activities. They are well known to oxidatively split carotenoids, which are colored compounds found abundantly in nature including in many foods. Beyond this oxidation reaction, certain CCDs catalyze other reactions involving carotenoids and their derivatives and also non-carotenoid molecules. These reactions are involved in several important biological processes including the production of universal light-sensing molecules, breakdown of spent plant matter, and generation of signaling molecules critical for plant and animal physiology. As such, control of CCD function has long been of interest in a variety of applied research fields including agriculture, horticulture, medicine, and biofuel production. Despite their importance, many critical gaps in knowledge remain regarding how CCDs achieve such varied activities, which consequently has limited our ability to control or modify their function. This research project seeks to provide a molecular level understanding of how CCDs achieve their diverse activities. The new knowledge may open new avenues for controlling the CCD activity using small molecule modulators or by rational alteration of their molecular structure. The participation of graduate and undergraduate students from underrepresented minority groups and those with military service in this research will help achieve a broader educational goal of promoting the entry of such individuals into STEM-fields.The primary focus of this research project is to elucidate the determinants of the varied activities that are exhibited by CCDs. The first aim is to uncover the mechanism by which CCDs trigger dioxygen reactivity using a variety of sophisticated spectroscopic techniques. The second aim will employ high-resolution X-ray crystallography to uncover the active site properties that allow certain CCDs to carry out trans-cis isomerization of carotenoid substrates. The third aim focuses on probing the breadth and mechanism of CCD substrate specificity by rational mutagenesis and the CCD activities found in archaea that occupy diverse ecological niches. Overall, this research will advance the understanding of the catalytic diversity of CCDs and will provide novel insights into their evolution and biological functions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

在化学系生命过程化学项目的支持下，加州大学欧文分校的 Philip Kiser 博士正在研究一种称为类胡萝卜素裂解双加氧酶 (CCD) 的酶家族。这些酶具有高度保守的 4-His 铁中心，并表现出非常多样化的催化活性。众所周知，它们会氧化分解类胡萝卜素，类胡萝卜素是一种在自然界（包括许多食物）中大量存在的有色化合物。除了这种氧化反应之外，某些 CCD 还催化涉及类胡萝卜素及其衍生物以及非类胡萝卜素分子的其他反应。这些反应涉及几个重要的生物过程，包括通用光敏分子的产生、废植物物质的分解以及对植物和动物生理学至关重要的信号分子的产生。因此，CCD 功能的控制长期以来一直受到农业、园艺、医学和生物燃料生产等各种应用研究领域的关注。尽管它们很重要，但关于 CCD 如何实现如此多样的活动，仍然存在许多关键的知识空白，这因此限制了我们控制或修改其功能的能力。该研究项目旨在从分子水平上了解 CCD 如何实现其不同的活性。新知识可能为使用小分子调节剂或通过合理改变其分子结构来控制 CCD 活性开辟新途径。来自代表性不足的少数群体和服兵役的研究生和本科生参与这项研究，将有助于实现促进这些人进入 STEM 领域的更广泛的教育目标。该研究项目的主要重点是阐明 CCD 所表现出的各种活动的决定因素。第一个目标是利用各种复杂的光谱技术揭示 CCD 触发双氧反应的机制。第二个目标将采用高分辨率 X 射线晶体学来揭示活性位点特性，从而允许某些 CCD 进行类胡萝卜素底物的反式-顺式异构化。第三个目标侧重于通过合理诱变探讨 CCD 底物特异性的广度和机制，以及在占据不同生态位的古细菌中发现的 CCD 活性。总体而言，这项研究将增进对 CCD 催化多样性的理解，并将为其进化和生物功能提供新颖的见解。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Preparation of carotenoid cleavage dioxygenases for X-ray crystallography

• DOI: 10.1016/bs.mie.2021.10.020
• 发表时间: 2022-01-01
• 期刊: CAROTENOIDS
• 作者: Daruwalla，Anahita;Sui，Xuewu;Kiser，Philip D.
• 通讯作者: Kiser，Philip D.

Chromophore hydrolysis and release from photoactivated rhodopsin in native membranes.

• DOI: 10.1073/pnas.2213911119
• 发表时间: 2022-11-08
• 期刊: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• 影响因子: 11.1
• 作者: Hong, John D.;Salom, David;Kubas, Adam;Kiser, Philip D.;Palczewski, Krzysztof
• 通讯作者: Palczewski, Krzysztof