Protein-Protein Interactions in Photosystem II

本项目旨在阐明高等植物光系统II相关蛋白的结构组织。光系统结构组分之间的蛋白质-蛋白质相互作用将使用生化和分子方法进行研究。首先，将研究锰稳定蛋白与24kda蛋白的相互作用。当锰稳定蛋白与PS II膜结合时，化学修饰会显著影响24 kDa蛋白与PS II结合的能力。它将确定24kda蛋白的结合抑制是由于电荷中和还是空间干扰。修饰残基的位置将通过质谱法绘制。其次，将确定化学修饰对24和17 kDa蛋白在蛋白质结合和功能方面的影响，并绘制修饰残基的位置。第三，观察到的PS II的外部亚单位的异质性和光系统的结构和功能的作用将被检查。在拟南芥中，PS II的每个外源蛋白都由两个基因编码。T-DNA插入系，可用于许多这些基因将被检查。对于没有T-DNA插入系的基因（psbO-2和psbP-1），将引入RNAi构建物来抑制这些基因产物的表达。这些植物的表型将在控制和胁迫条件下对PS II的功能、组装和稳定性进行研究。除了这种遗传方法，生化重建研究检查两个psbO蛋白的功能将被执行。这些研究将为PS II的外在成分和光系统的内在成分以及彼此之间的相互作用提供重要的新信息。此外，对外在亚基异质性作用的理解可能为光合作用的调控“微调”提供见解。最后，拟南芥突变体，我们将产生将提供一个资源来检查外部亚单位在体内的作用。更广泛的影响：该项目将涉及不同层次的培训。预计将有几名本科生参与与该项目相关的研究。这些学生将参加科学会议，展示他们的研究并最终发表他们的成果。研究生、博士后研究人员和研究助理都密切参与这些本科生研究人员的培训。此外，该项目还将继续与东卡罗莱纳大学（East Carolina University）的女教员辛迪·普特南-埃文斯（Cindy Putnam-Evans）博士进行长期合作。发放部门。

This project will elucidate the structural organization of proteins associated with Photosystem II (PS II) in higher plants. Protein-protein interactions among the structural components of the photosystem will be studied using biochemical and molecular approaches. First, the interaction of the manganese-stabilizing protein with the 24 kDa protein will be examined. Chemical modification of the manganese-stabilizing protein when it is associated with PS II membranes dramatically affects the ability of the 24 kDa protein to bind to PS II. It will determined if the binding inhibition of the 24 kDa protein is due to charge neutralization or steric interference. The location of the modified residues will be mapped by mass spectrometry. Second, the effects of chemical modification on the 24 and 17 kDa proteins with respect to protein binding and function will be determined and the location of the modified residues will be mapped. Third, the role of the observed heterogeneity in the extrinsic subunits of PS II with respect to structure and function of the photosystem will be examined. In Arabidopsis, each of the extrinsic proteins of PS II are encoded by two genes. T-DNA insertion lines, which are available for a number of these genes will be examined. For genes, which have no T-DNA insertional lines available (psbO-2 and psbP-1), RNAi constructs will be introduced to suppress the expression of these gene products. The phenotypes of these plants will be examined with respect to PS II function, assembly and stability under control and stress conditions. In addition to this genetic approach, biochemical reconstitution studies examining the function of the two psbO proteins will be performed. These studies will provide important new information concerning the interactions of the extrinsic components of PS II with the intrinsic components of the photosystem and with each other. Additionally, an understanding of the role of extrinsic subunit heterogeneity may provide insights into regulatory "fine tuning" of the photosynthesis. Finally, the Arabidopsis mutants, which we will generate will provide a resource for examining the role of the extrinsic subunits in vivo.Broader Impacts: This project will involve training at different levels. It is anticipated that several undergraduate students will be involved in research related to this project. These students will attend scientific meetings, presenting their research and ultimately publish their results. The graduate students, postdoctoral researchers and research associates are all intimately involved in the training of these undergraduate researchers. Additionally, the project will continue a long-standing association with Dr. Cindy Putnam-Evans at East Carolina University, a female faculty member in a non-Ph.D.-granting department.