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The Tangled Signaling Pathway of a Bacterial Phytochrome

细菌光敏色素的纠结信号通路

基本信息

批准号：
1518160
负责人：
Katrina Forest
金额：
$ 65万
依托单位：
University of Wisconsin-Madison
依托单位国家：
美国
项目类别：
Standard Grant
财政年份：
2015
资助国家：
美国
起止时间：
2015-08-01 至 2019-07-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=1518160&HistoricalAwards=false
关键词：
Tangled Signaling Pathway Bacterial Phytochrome

项目摘要

Phytochromes are light-responsive sensory molecules, but exactly how light is perceived by bacteria and how that information is used to bring about adaptation to changes in light levels is not well understood. This project will explore how protein folding influences the signaling pathways of a soluble phytochrome. Additionally, this project will investigate how other proteins (called chaperones) facilitate the protein folding required for proper phytochrome activity. These experiments promise to provide generally applicable insights into phytochromes, and in addition phytochromes will be modified to create new microscopic imaging tools which will permit biologists to peer into living organisms to learn more about basic biological processes. The project includes outreach activities that facilitate interaction between scientists and artists to enhance science communication and learning. The objective of this project is to understand how the unusual knotted topology of bacterial phytochrome determines the signaling properties and folding pathway of this light-regulated histidine kinase. Phytochrome is one of few deeply knotted proteins known, and the route to its figure-of-eight final topology is completely unknown. To determine whether the knotted protein structure of phytochrome and the catenated dimeric interface of its cognate response regulator contribute to intra- and intermolecular signaling, the project will utilize kinase, phosphotransferase, and phosphatase assays on native and untangled (engineered) bacterial phytochromes and their cognate response regulators. To determine how the folding pathway of knotted phytochrome is dependent on characteristics of the proline-rich lasso loop the project will investigate the effects of sequence and length variation in this loop on folding of phytochromes in the presence and absence of a prolyl-isomerase containing chaperone. To create small genetically encoded fluorophores with desirable near infrared excitation maxima, the project will reverse engineer phytochrome by removing domains required only for signal transduction while maintaining elements required for stable folding and chromophore interaction.

光敏色素是对光敏感的感觉分子，但细菌究竟如何感知光线，以及如何利用这些信息来适应光线水平的变化，目前还没有很好的了解。这个项目将探讨蛋白质折叠如何影响可溶性光敏色素的信号通路。此外，该项目还将研究其他蛋白质（称为伴侣蛋白）如何促进适当光敏色素活性所需的蛋白质折叠。这些实验有望为光敏色素提供普遍适用的见解，此外，光敏色素将被修改，以创建新的显微成像工具，这将使生物学家能够窥视活生物体，以了解更多关于基本生物过程。该项目包括促进科学家和艺术家之间互动的外联活动，以加强科学传播和学习。本项目的目的是了解细菌光敏色素的不寻常的打结拓扑结构如何决定这种光调节组氨酸激酶的信号传导特性和折叠途径。光敏色素是已知的少数几种深度打结的蛋白质之一，其最终8字形拓扑结构的路线完全未知。为了确定光敏色素的打结蛋白质结构及其同源反应调节剂的链状二聚体界面是否有助于分子内和分子间信号传导，该项目将利用天然和未缠结（工程化）细菌光敏色素及其同源反应调节剂的激酶，磷酸转移酶和磷酸酶测定。为了确定打结光敏色素的折叠途径如何依赖于富含脯氨酸的套索环的特征，该项目将研究在存在和不存在脯氨酰异构酶的情况下，该环中的序列和长度变化对光敏色素折叠的影响。为了创造具有理想的近红外激发最大值的小的遗传编码荧光团，该项目将通过去除仅用于信号转导的结构域，同时保留稳定折叠和发色团相互作用所需的元素，对光敏色素进行逆向工程。