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

1)用改进的方法制备牛心脏线粒体复合体III(BC_1)。通过稳态反应动力学分析，该晶体在将电子从QH_2转移到细胞色素c2的过程中表现出乒乓机制。2)在室温下，通过含有蔗糖的聚乙二醇化处理，该BC_1络合物可在1-2周内大规模结晶成红色平行管状。3)该晶体在低于8A的分辨率下对X射线进行了衍射，但在机械冲击下相当脆弱，暴露在空气中不稳定。4)如果以还原的形式制备Euglena细胞色素c，则其稳定性较好，为结晶提供了良好的准备。5)完成了Euglena细胞色素c_1编码基因的碱基序列。不同C_1序列的比较表明，His和Met是血红素铁原子的配体。6)酵母细胞色素c中可能的血红素配体His和Met的定点突变表明，这两个配体对BC_1复合体的铁络合和组装具有重要作用。7)通过基因缺失实验发现，酵母细胞色素c_1中70位左右与细胞色素c相互作用的两个酸性区域中的一个对相互作用不是必需的。8)13 kDa(两种不同的多肽)、20kDa、9)蓝藻b_6f复合体能够通过细胞色素c&lt将电子传递给细胞色素AA_3；553&gt；

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>.