Controlling Enzyme Structure and Activity with Light Illumination

用光照控制酶的结构和活性

• 批准号: 0554115
• 负责人: C. Ted Lee, Jr.
• 金额: --
• 依托单位: University of Southern California
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0554115&HistoricalAwards=false
• 关键词: Controlling; Enzyme; Structure; Activity; Light

AbstractProposal Title: Controlling Enzyme Structure and Activity with Light Illumination Proposal Number: CTS-0554115Principal Investigator: C. Ted Lee, Jr.Institution: University of Southern CaliforniaAnalysis (rationale for decision): Enzymes interact with a variety of molecules during the course of activity, resulting in conformation changes to intermediately-folded states that influence nearly all biological processes. Thus, the general form-function relationships of proteins should not be considered static; instead, to fully understand this relationship requires knowledge of how a protein or enzyme unfolds in response to various molecules. To properly investigate this phenomenon requires two complementary approaches: (1) a means to induce protein unfolding in a controlled manner, and (2) a means to study the conformation of unfolded proteins at relatively high resolutions. To achieve this tandem goal, two novel techniques will be pursued in this project, namely the interaction of proteins with photoreversible surfactants, and the use of small-angle neutron scattering (SANS) to determine the structure of partially-folded/unfolded enzymes in response to photoresponsive surfactants. Surfactants unfold proteins by disrupting intramolecular amino acid contacts. In the case of photoresponsive surfactants, the surfactant-amino acid associations can be switched on or off with simple light illumination, resulting in photocontrol of protein conformation. Protein conformation will then be determined with SANS experiments by applying shape-reconstruction methods to optimize the positions of scattering centers (i.e., protein amino acids or individual atoms, depending on the desired resolution) in order to best fit the scattering data. This combined approach will allow full characterization of the photocontrolled native intermediate denatured transitions in enzymes and proteins, providing unique insight into folding pathways. By controlling protein form with light, photocontrol of protein function will also be achieved, allowing enzymatic activity to be switched on and off with light illumination.Photo-controlled enzyme folding requires at a minimum nothing more than a common, commercial black light and, as such, can be easily ported to the classroom to provide an uncomplicated protocol to seamlessly blend research into coursework. This will serve to reconnect the flow of ideas from the laboratory to the curriculum, and will better prepare students to meet the challenges of a technically diverse, modern day workplace. These efforts will be disseminated to the scientific community, particularly the results of Degree Projects, four-year research projects that provide horizontal (over-time) and vertical (cross-course) integration of research and education.

摘要建议标题：用光照明控制酶的结构和活性建议编号：CTS-0554115主要研究人员：C.Ted Lee，Jr.机构：南加州大学分析(决策的基本原理)：酶在活动过程中与各种分子相互作用，导致构象变化到中间折叠状态，几乎影响所有生物过程。因此，蛋白质的一般形式-功能关系不应被认为是静态的；相反，要完全理解这种关系，需要了解蛋白质或酶如何对各种分子做出反应。为了正确地研究这一现象，需要两种互补的方法：(1)以受控方式诱导蛋白质去折叠的方法；(2)以相对较高的分辨率研究未折叠蛋白质的构象的方法。为了实现这一目标，该项目将采用两种新的技术，即蛋白质与光可逆表面活性剂的相互作用，以及利用小角中子散射(SANS)来确定部分折叠/未折叠的酶对光响应性表面活性剂的响应结构。表面活性剂通过破坏分子内氨基酸的接触来展开蛋白质。在光响应型表面活性剂的情况下，表面活性剂-氨基酸的缔合可以在简单的光照射下打开或关闭，从而导致对蛋白质构象的光控制。然后将通过SANS实验确定蛋白质构象，方法是应用形状重建方法来优化散射中心(即蛋白质氨基酸或单个原子，取决于所需的分辨率)的位置，以便最好地拟合散射数据。这种结合的方法将允许充分表征光控制的酶和蛋白质中的天然中间变性转变，为折叠途径提供独特的见解。通过用光控制蛋白质的形式，还可以实现对蛋白质功能的光控制，允许在光照明下打开和关闭酶的活性。光控制的酶折叠至少只需要一个普通的商业黑灯，因此可以很容易地移植到教室，提供一个简单的协议，将研究无缝地融入到课程中。这将有助于将从实验室到课程的思想流动重新联系起来，并将使学生更好地准备迎接技术多样化的现代工作场所的挑战。这些努力将向科学界传播，特别是学位项目的成果，即提供横向(长期)和纵向(跨学科)研究与教育一体化的四年期研究项目。