Collaborative Research: Investigations of the Plant Photoreceptor Kinase Family of Phototropins

植物光感受器介导从生理到生化和分子水平的广泛的光反应，并存在于植物发育的整个过程，从种子萌发到几乎所有的营养生长阶段，直到开花和衰老。目前已知的光感受器有光敏色素、隐色素和趋光素。促光体激素代表了一类新的、以前未被描述过的光感受器。这些双发色光受体结合了FMN的两个分子，不同于已知的任何一类光感受器，它们通过独特的初始光化学作用并导致自动磷酸化。本项目的重点是嗜铬蛋白促红细胞生成素(Nph1)和迄今为止所描述的各种nph1样蛋白。这些蛋白质在异源系统中以发色团结合域和全长蛋白质的形式表达。表达的蛋白质，包括天然结构和定点突变的蛋白质，都是以光活性形式存在的。我们计划对各种分离的生色团结合域和全长蛋白质进行有限的生化和详细的生物物理表征。生化研究将强调使用突变结构来探索初始光化学。后者将专注于探索光激活光化学的分子机制及其在蛋白质水平上的后果。这些研究将涉及时间分辨光学和振动光谱学。生色团结合域与从古生物到哺乳动物的各种蛋白质中的结构域(所谓的PAS结构域)密切相关，应该为我们理解这些结构域的功能提供一个新的维度。

0090837BogomolniPlant photoreceptors mediate a broad spectrum of responses to light, ranging from the physiological through to the biochemical and molecular levels, and are present throughout plant development from seed germination through virtually all phases of vegetative growth to flowering and senescence. Currently well-known photoreceptors are phytochromes, cryptochromes, and phototropins. Phototropins represent a new and previously uncharacterized class of photoreceptors. These dual chromophoric photoreceptors, binding two molecules of FMN, are unlike any of the known classes of photoreceptors , and operate through an initial photochemistry that is unique and leads to autophosphorylation. The present project is focused on the chromoprotein phototropin (nph1) and the various nph1-like proteins described to date. These proteins are expressed in heterologous systems as the chromophore-binding domains and as the full length proteins. The expressed proteins both the native constructs and those with site-specific mutations are in the photoactive form. We plan to undertake a limited biochemical and detailed biophysical characterization of the various isolated chromophore-binding domains and of the full-length protein. The biochemical studies will emphasize the use of mutant constructs to probe the initial photochemistry. The latter will focus on studies probing the molecular mechanism by which light activates the photochemistry and its consequences at the protein level. These studies will involve time-resolved optical and vibrational spectroscopy. The chromophore-binding domains are closely related to domains (so-called PAS domains) in a diverse range of proteins from archaea through mammals, and should provide a new dimension to our understanding of the functions of these domains.