Mutual Recognition and Electron Transfer of Electron Carriers: Cytochrome and Ferredoxin Systems

牛心脏线粒体细胞色素<C_1>亚复合物与3mol / l的pCMB反应修饰3个sh -基团。这两个亚基（血红素亚基和铰链蛋白）没有通过凝胶过滤分离，而是通过电泳分离，导致<C_1>血红素亚基出现自氧化性。这表明铰链蛋白不仅在<C_1>和c之间形成复合物，而且在阻碍<C_1>血红素亚基自氧化方面是必不可少的。如果铰链蛋白不相关，血红素亚基似乎会聚合。将细胞色素<C_1>的一些突变基因导入酵母细胞，观察其对酵母生长、呼吸能力、光谱特性、线粒体内膜组装和基因表达的影响。从<C_1>的羧基末端区域删除17个氨基酸对这些特性没有影响，但当从羧基末端区域删除70个氨基酸时，修饰蛋白失去了融入呼吸链复合体III的能力，失去了生长和呼吸的能力。这在建立牛肉细胞色素<C_1>结构时得到了部分预测。这条线上的工作不仅对阐明细胞色素<C_1>的结构-功能关系，而且对阐明其他膜蛋白的结构-功能关系具有重要的意义。本学期完成的其他工作包括：血红素铁配体的鉴定，<C_1>和铁氧还蛋白<NADP^+>还原酶的结晶，该酶中sh -基团的表征，铁硫簇的形成。

The cytochrome <C_1> subcomplex of beef heart mitochondria was reacted with 3 molar equivalents of pCMB to modify 3 SH-groups. The two subunits (heme-subunit and hinge protein) were not separated on gel filtration, but by electrophoresis after this treatment, leading to the appearance of autoxidizability of <C_1> heme subunit. This indicates that the hinge protein is indispensable not only for complex formation between <C_1> and c, but also for obstructing the <C_1> heme subunit from autoxidation. The heme subunit seems to polymerize if the hinge protein is not associated.Some mutated genes of cytochrome <C_1> were introduced into yeast cells to see how they affect the growth of yeast, respiratory ability, spectral characteristics, assembly into mitochondrial inner membrane, and gene expression. The deletion of 17 amino acids from the carboxyl-terminal region of <C_1> did not affect at all these characteristics, but when 70 amino acids from the carboxyl-terminal region were deleted, the protein modified lost the ability to be integrated into complex III of respiratory chain, and growth as well as respiration. This was partly predicted when the structure of beef cytochrome <C_1> was established. The work on this line is very promising to elucidate structure-function relationship of not only cytochrome <C_1> , but other membranous proteins. Other works done during this term are as follows: Identification of the heme iron ligands, crystallizations of <C_1> and ferredoxin <NADP^+> reductase, characterization of SH-groups in this enzyme, and iron-sulfur cluster formation.