Structure-Function Studies on Phytochrome

这些研究的总体目标是确定光敏色素光感受器的光感觉和调节功能的结构基础。这些研究依赖于表达、组装和纯化重组光敏色素的能力。以往的生物化学和遗传学研究将光敏色素分子划分为两个主要结构域，一个是位于分子N端高度保守的光感结构域，另一个是在光敏色素的调控功能中起关键作用的c端结构域。这两个领域是这些研究的重点。第1部分概述的实验旨在鉴定光敏色素光敏区域中对发色团-蛋白质相互作用很重要的特定氨基酸残基。这些研究将利用化学交联、定点和随机诱变以及共振拉曼光谱技术。这些研究也代表了光敏色素的x射线晶体学分析的前奏（和辅助）。第二部分概述的实验解决了光敏色素作为光调节酶的假设。令人兴奋的是，在蓝藻中发现了与高等植物光敏色素和原核生物的递质蛋白激酶家族在调控区域内共享结构基元的光敏色素相关基因，为验证这一有争议的理论提供了新的动力。实验试图确定植物*-铬是否是一种光调节的递质激酶，如果是，则确定其酶活性的结构基础。本节还概述了真核生物和原核生物光敏色素之间生化和功能“交叉对话”的平行研究，将为光敏色素结构和功能的进化提供见解。适应外部光环境变化的能力对植物的生存至关重要，因为与动物不同，植物必须有阳光才能进行光合作用，它们缺乏向更理想的光环境移动的能力。因此，植物拥有光敏分子，使它们能够适应环境中的次优光照条件，其中一种被称为光敏色素。光敏色素介导的许多现象包括诱导种子萌发，响应邻近植物遮荫的茎快速伸长，调节开花时间和向光生长。光敏色素是一种蛋白质，它含有一种附着的色素，这种色素负责吸收可见光。光敏色素在吸收光后转化为一种调节植物生长发育过程的活性物质。这项工作旨在剖析光敏色素分子的解剖结构，并描述使其能够将光信号转换为化学信号的结构特征。更好地了解光敏色素的作用机制将为改善植物的生长发育提供新的途径。

9604511 Lagarias The overall goal of these studies is to define the structural basis for photosensory and regulatory functions of the phytochrome photoreceptor. These investigations rely on the ability to express, assemble and purify recombinant phytochromes. Previous biochemical and genetic studies have delimited the phytochrome molecule into two major domains, a highly conserved photosensory domain at the molecule's N terminus and a C-terminal domain which plays a key role in the regulatory function of phytochrome. These two domains are the focus of these studies. Experiments outlined in Part 1 seek to identify specific amino acid residues in the photosensory domain of phytochrome that are important to chromophore-protein interactions. These studies will exploit the techniques of chemical cross-linking, site-directed and random mutagenesis, and resonance Raman spectroscopy. These studies also represent a prelude (and adjunct) to x-ray crystallographic analysis of phytochrome. Experiments outlined in Part II address the hypothesis that phytochrome functions as a light-regulated enzyme. The exciting discovery of phytochrome-related genes in cyanobacteria, which share structural motifs with higher plant phytochromes and the transmitter protein kinase family of prokaryotes within the regulatory domain region, has provided renewed impetus for testing this controversial theory. Experiments seek to establish whether phyto*-chrome is a light-regulated transmitter kinase and if so, to define the structural basis for its enzymatic activity. Parallel studies to test for biochemical and functional `cross-talk' between eukaryotic and prokaryotic phytochromes, also outlined in this section, will provide insight into the evolution of phytochrome structure and function. The ability to adjust to changes in their external light environment is essential to plant survival because, unlike animals, plants must have sunlight for photosynthesis and they lack the ability to move to a more optimum light environment. Plants therefore possess light-sensing molecules which enable them to adapt to suboptimal light conditions in their environment, one of which is called phytochrome. Among the many phenomena mediated by phytochrome include the induction of seed germination, rapid stem elongation in response to the shade of neighboring plants, regulation of the timing of flowering and growth towards light. Phytochrome is a protein that contains an attached pigment which is responsible for its ability to absorb visible light. Upon light absorption, phytochrome is converted to an active form which regulates the course of plant growth and development. This work seeks to dissect the anatomy of the phytochrome molecule and to characterize the structural features that enable it to convert a light signal into a chemical signal. A better understanding of the mechanism of phytochrome action should lead to novel approaches to improve plant growth and development.