Structural Organization of Proteins on the Oxidizing Side of Photosystem II

光系统 II 氧化侧蛋白质的结构组织

• 批准号: 9006552
• 负责人: Terry Bricker
• 金额: $ 26.3万
• 依托单位: Louisiana State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-08-01 至 1994-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9006552&HistoricalAwards=false
• 关键词: Structural; Organization; Proteins; Oxidizing; Side

This project describes the production and preliminary characterization of mutants bearing lesions in the extrinsic loop region of CPal in the cyanobacterium Synechocystis 6803. CPal (encoded by the psbB gene) is an intrinsic membrane protein which is an integral component of the photosynthetic apparatus of cyanobacteria and higher plants and appears to be required for photosynthetic oxygen evolution in Photosystem II. The goal of this project is to determine the function of the large and unusual extrinsic loop region of this protein. This region appears to interact with the extrinsic 33 kDa protein and possibly the inorganic cofactors manganese, chloride, or calcium, of all of which are required for oxygen evolution. Additionally, CPal appears to act as a chlorophyll-a antenna for Photosystem II. The extrinsic loop region may interact with other antennae proteins and/or the Photosystem II reaction center. In vitro mutagenesis (saturation and oligonucleotide-mediated site- directed) will be used to produce mutations in the extrinsic loop region. These mutations will be re-introduced into Synechocystis (along with a closely linked antibiotic resistance marker), and mutants will be isolated by a combination of screening for antibiotic resistance and loss of photoautotrophic growth. Additionally, screening for antibiotic resistance and loss of photoautotrophic growth. Additionally, screening for temperature sensitive mutations will be performed. These mutations will then be characterized by examination of their electron transport activity 77oK fluorescence emission spectra, manganese and calcium content of Photosystem II-enriched preparations, photosynthetic membrane protein composition, and their chloride requirements for the stabilization of manganese.

该项目描述了蓝藻集胞藻 6803 CPal 外环区域中带有损伤的突变体的产生和初步表征。CPal（由 psbB 基因编码）是一种内在膜蛋白，是蓝藻和高等植物光合装置的组成部分，似乎是光合作用氧气所必需的 光系统 II 的进化。 该项目的目标是确定该蛋白质的大且不寻常的外在环区域的功能。 该区域似乎与外在 33 kDa 蛋白质相互作用，并且可能与无机辅助因子锰、氯化物或钙相互作用，所有这些都是氧气释放所必需的。 此外，CPal 似乎充当光系统 II 的叶绿素天线。 外在环区域可能与其他触角蛋白和/或光系统 II 反应中心相互作用。 体外诱变（饱和和寡核苷酸介导的定点）将用于在外在环区域中产生突变。 这些突变将被重新引入集胞藻（以及紧密相连的抗生素抗性标记），并且将通过抗生素抗性筛选和光合自养生长丧失的结合来分离突变体。 此外，筛查抗生素耐药性和光合自养生长丧失。 此外，还将进行温度敏感突变的筛查。 然后，将通过检查其电子传递活性 77oK 荧光发射光谱、富含光系统 II 的制剂中的锰和钙含量、光合膜蛋白组成以及稳定锰的氯化物需求来表征这些突变。