Structure, Function, and Evolution of Eukaryotic and Prokaryotic Respiratory Electron Transfer Systems

真核和原核呼吸电子传递系统的结构、功能和进化

基本信息

批准号：
01470148
负责人：
MATSUBARA Hiroshi
金额：
$ 4.1万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1989
资助国家：
日本
起止时间：
1989 至 1990
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-01470148/
关键词：
Respiratory Electron Transfer Systems Prokaryote And Eukaryote Structure, Function And Evolution Cytochrome bc_1 Complex Crystallization Of bc_1 Site Directed Mutagnesis Euglena Cytochrome c_1 08Cyanobacterial Cytochrome b_6f ウシ心筋チトクロムbc_1

项目摘要

1) Bovine heart mitochondrial complex III (bc_1 was prepared by an improved method. This preparation showed a ping-pong mechanism analyzed by a steady-state reaction kinetics, when it transfers electrons from QH_2 to cytochrome c.2) This bc_1 complex was reproducibly crystallized in a large scale to reddish parallelpipe forms by treating the preparation with polyethyleneglycol containing sucrose within 1-2 weeks at room temperature.3) The crystal diffracted X-ray at less than 8 A resolution, but it was rather fragile for mechanical shocks and unstable by exposing it to air. Therefore, some improvements in crystallization and manipulations to X-ray analysis.4) Euglena cytochrome c was found to be stable if it was prepared in the reduced form, promisingit to be a good preparation for crystallization.5) The base sequence of cDNA encoding Euglena cytochrome c_1 was completed. The comparison of various c_1 sequences revealed that His and Met were the ligands to the heme iron atom. Heme C was bound to only one cysteine through a thioether bond and Phe was in the position of Cys usually binding the heme together with the other.6) The site directed mutations of putative heme ligands, His and Met, in yeast cytochrome c_1 showed that these two have important roles for the iron chelation and assembly of the bc_1 complex.7) One of the two acidic regions supposed to interact with cytochrome c at around position 70 in yeast cytochrome c_1 was found to be not essential for the interaction by gene deletion experiments.8) Sequence analyses of 13kDa(two different polypeptides), 2okDa, and 20kDa Sub-units revealed that there are several regions forming iron-sulfur clusters in mitochondrial Complex I.9) A cyanobacterial b_6f complex was found to be able to transfer electrons to cytochrome aa_3 through cytochrome c_<553>.

1）牛心脏线粒体复合物III（BC_1是通过改进的方法制备的。这种制备显示通过稳态反应动力学分析的乒乓机制，当它从QH_2转移到cytochrome c.2时，该bc_1复合物在大尺度上以重新构造的形式将电子转移到ch_2。 3）晶体衍射的X射线在少于8的分辨率下进行蔗糖，但是对于机械冲击而言，它相当脆弱，并且通过将其暴露于空气而不稳定。因此，对X射线分析的结晶和操作的一些改进。4）Euglena细胞色素C如果以还原形式制备，则发现它是结晶的良好准备。5）编码Euglena CytoChrome C_1的CDNA的基础序列。各种C_1序列的比较表明，他和MET是血红素铁原子的配体。血红素C仅通过硫醇键和PHE结合到一个半胱氨酸，通常在Cys位置通常与另一个半胱氨酸与另一个半胱氨酸结合。6）6）该地点在酵母菌C_1中指示他和MET的位点，在酵母菌C_1中，这两个位置表明，这两种表明这两个与BC_1复杂的铁螯合物和组件相互作用。发现酵母菌C_1中的位置在70位的位置在基因缺失实验中的相互作用中并不是必不可少的。8）13KDA的序列分析（两个不同的多肽），2OKDA和20KDA子单位，以及20KDA子单位的序列分析表明，有几个区域在MiTochrial Complectrial I.9中成立了几个区域，该区域是MiTochrial compliprial i.9）cyan cyanobs to cyan of cyanobs to cyan cynob to cyan cynob i.9）通过细胞色素C_ <553>电子到细胞色素的电子AA_3。