Functional expression and utilization of bacterial cytochrome C__-

细菌细胞色素C__-的功能表达和利用

• 批准号: 03556010
• 负责人: KODAMA Tohru
• 金额: $ 10.11万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1991
• 资助国家: 日本
• 起止时间: 1991 至 1992
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-03556010/
• 关键词: cytochrome C__-; electro chemistry; シトロクロムC; 電気化学反応

Cytochrome c__-_<552> gene was cloned from the total DNA of the cells of Hydrogen obacter thermophilus. The gene was shown to contain a signal sequence. As a next step, the gene without the signal sequence was prepared. When Escherichia coli was transformed with the plasmid containing this gene and was cultivated under nitrate respiration, it produced cytochrome c__-_<552> within cytoplasm fraction.Cytochrome c__-_<551> gene was also cloned from the total DNA of the cells of Pseudomonas aeruginosa. When Pseudomonas aeruginosa was transformed with the plasmid containig this gene, it produced cytochrome c__-_<551> within periplasm fraction. The production rate was as high as 0.7 % of the total protein of the cell-free extract. An attempt was made to produce cytochrome c__-_<551> by using the other microorganisms (Escherichia coli or Pseudomonas putida), however, successful production could not be obtained.Reactivity of the two proteins (cytochrome c__-_<552> and cytochrome c__-_<551>) toward electrode was also investigated. Both cytochromes were shown to react with Pt-electrode in a solution state. Cytochrome c__-_<552> protein could react with Au-electrode in the presence of promotor, however, in its absence reactivity was not observed.

细胞色素c__-_<552>基因是从嗜热氢杆菌细胞的总DNA中克隆的。该基因被证明含有信号序列。下一步，制备没有信号序列的基因。当用含有该基因的质粒转化大肠杆菌并在硝酸盐呼吸下培养时，它在细胞质部分内产生细胞色素c__-_<552>。细胞色素c__-_<551>基因也从铜绿假单胞菌细胞的总DNA中克隆。当用含有该基因的质粒转化铜绿假单胞菌时，它在周质部分内产生细胞色素c__-_<551>。无细胞提取物总蛋白的产率高达0.7%。尝试使用其他微生物（大肠杆菌或恶臭假单胞菌）生产细胞色素c__-_<551>，但未能成功生产。还研究了两种蛋白质（细胞色素c__-_<552>和细胞色素c__-_<551>）对电极的反应性。两种细胞色素均在溶液状态下与铂电极发生反应。在启动子存在的情况下，细胞色素 c__-_<552> 蛋白可以与金电极反应，然而，在启动子不存在的情况下，没有观察到反应性。

项目成果

Arai,H.et al.: "Anaerobically Induced Expression of the Nitrite Reductase Cytochrome c-551 Operon from Pseudomonas aeruginosa" FEBS Letters. 280. 351-353 (1991)

Arai,H.et al.：“来自铜绿假单胞菌的亚硝酸盐还原酶细胞色素 c-551 操纵子的厌氧诱导表达”FEBS 快报。

Hiroyuki Arai: "Nitrite actives the transcription of the Pseudomonas aeruginosa nitrite reductase and cytochrome c__-_<551> under anaerobic conditions" FEBS Letters. 288. 227-228 (1991)

Hiroyuki Arai：“亚硝酸盐在厌氧条件下激活铜绿假单胞菌亚硝酸还原酶和细胞色素 c__-_<551> 的转录” FEBS Letters。

Hiroyuki ARAI: "Nitrite activates the transcription of the Pseudomonas aeruginosa nitrite reductase and cytochrome C_<551> under anaerobic conditions" FEBS Letters. 288. 227-228 (1991)

Hiroyuki ARAI：“亚硝酸盐在厌氧条件下激活铜绿假单胞菌亚硝酸还原酶和细胞色素 C_<551> 的转录”FEBS Letters。