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Expression of ferredoxin gene family and its physiological significance

铁氧还蛋白基因家族的表达及其生理意义

基本信息

批准号：
02660086
负责人：
HASE Toshiharu
金额：
$ 1.28万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02660086/
关键词：
Maize ferrecoxin photosynthesis gene cloning glutamate synthase glutamine synthetase 遺伝子発現光誘導

项目摘要

Maize (Zea mays L.) has two types of ferredoxin (Fd) differentially expressed in photosynthetic and non-photosynthetic organs. To understand structural characteristics of the isoproteins and molecular mechanism of the differential expression of their genes, we have cloned two different maize cDNAs, which encode the entire precursor polypeptides of Fd I and Fd III composed of 150 and 152 amino acid residues, respectively. The gene transcript for Fd I was restricted to leaves and their levels increased markedly upon illumination, whereas that for Fd III was detected in all organs and its accumulation was not light dependent.To clarify the biosynsthetic pathway of the Fds present in different organs, in vitro synthesized precursors were examined to be imported into chloroplasts and etioplasts. Both Fds were processed to the mature size and assembled with the Fe-S cluster to become a functional molecule. The cluster assembly occured without tight coupling to the translocation, and the process was time and temperature dependent and did not require light.A cDNA fragment encoding the mature polypeptide of Fd III was expressed in E. coli, and the Fd was synthesized as a holo-form assembled with the cluster. Comparison of electron transfer activity of the two types of Fd showed that Fd III was superior as an electron acceptor from NADPH, and Fd I was superior as an electron donor for NADP, in reactions catalyzed by Fd-NADP reductase from maize leaf. The CD spectra of the two Fds also indicated a subtle difference in the geometry of their iron-sulfur clusters. These results are consistent with the view that photosynthetic and non-photosynthetic Fds may be functionally differentiated.

玉米（Zea mays L.）有两种类型的铁氧还蛋白（Fd）在光合和非光合器官中差异表达。为了了解同工蛋白的结构特征及其基因差异表达的分子机制，我们克隆了两种不同的玉米cdna，它们分别编码Fd I和Fd III的全部前体多肽，分别由150和152个氨基酸残基组成。Fd - 1基因转录本仅局限于叶片，光照条件下其表达量显著增加，而Fd - 3基因转录本在所有器官中均有表达，且其积累不依赖于光。为了阐明Fds在不同器官中的生物合成途径，研究了体外合成的前体导入叶绿体和病变体的情况。这两个Fds都被加工到成熟的尺寸，并与Fe-S簇组装成为功能分子。簇的组装与易位没有紧密耦合，并且该过程依赖于时间和温度，不需要光。在大肠杆菌中表达了一个编码Fd III成熟多肽的cDNA片段，并合成了与簇组装的Fd。在玉米叶片Fd-NADP还原酶催化的反应中，两种类型Fd的电子转移活性比较表明，Fd III作为NADPH的电子受体更优，Fd I作为NADP的电子给体更优。两个Fds的CD光谱也表明它们的铁硫团簇的几何形状有细微的差异。这些结果与光合和非光合Fds可能存在功能分化的观点一致。