Analysis of nucleoid proteins in tobacco chloroplasts

烟草叶绿体中核蛋白的分析

• 批准号: 02660092
• 负责人: SATO Fumiko
• 金额: $ 1.28万
• 依托单位: Kyoto 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-02660092/
• 关键词: Chloroplast; Nucleoids; Cultured green cells; DNA-binding protein; N-terminal aminoacid; Southwestern hybridization; Cholroplast differentiation; Transcription; 葉緑体核様体; 緑葉; 形質転換; rbcs遺伝子

Chloroplasts are cellular organelles, specific to plant cells and one of the differentiated form of plastids. While the genome structure, polypeptides and transcription regulation in chloroplasts have been investigated intensively, the controlling mechanism of chloroplast differentiation is still not clear. Here, we characterized the DNA binding proteins in chloroplast nucleoid(DNA-protein complex)of cultured tobacco(Nicotiana tabacum cv. Samsun NN)cells to get some information about the controlling mechanism of chloroplast differentiation. Nucleoids were purified by sequential density gradient centrifugation using metrizamid from purified chloroplasts. Electron microscopic observation of the purified nucleoid fraction and the treatment with proteinase or 2M NaCl indicated that nucleoids consist of DNAs and proteins. Southwestern hybridization with 32P-labeled chloroplast DNA indicated that some polypeptides in nucleoid fraction could bind DNA. The molecular weight of a polypeptide with DNA binding affinity isolated from cultured tobacco cells was about 4lkDa in denatured form. Competition experiment in southwestern hybridization with salmon DNA suggested that DNA binding was rather specific to cholroplast DNA. Determination on N-terminal amino acid sequence of 41kD DNA-binding polypeptide indicated that this polypeptide was not coded by choloroplast geneme.

叶绿体是细胞细胞器，特定于植物细胞和质体的分化形式之一。尽管已经对叶绿体中的基因组结构，多肽和转录调节进行了深入研究，但叶绿体分化的控制机制仍然不清楚。在这里，我们表征了培养的烟草（烟草taba​​cumcv。samsunnn）细胞中叶绿体核苷（DNA-蛋白复合物）中的DNA结合蛋白，以获取有关叶绿体分化的控制机制的一些信息。使用纯化的叶绿体的metrizamid通过顺序密度梯度离心纯化核苷。电子显微镜观察纯化的核苷馏分和用蛋白酶或2M NaCl处理的核苷酸是由DNA和蛋白质组成的。与32p标记的叶绿体DNA杂交西南杂交表明，核苷分数中的某些多肽可以结合DNA。从培养的烟草细胞中分离出的DNA结合亲和力的多肽的分子量约为4LKDA。与鲑鱼DNA的西南杂交中的竞争实验表明，DNA结合相当特定于胆红性DNA。对41KD DNA结合多肽的N末端氨基酸序列的测定表明该多肽未用胆管塑料基因编码。

项目成果

S.Takeda et al.: "Characterization of polypeptides that accumulate in cultured Nicotiana tabacum cells" Plant Cell Physiol.31. 215-221 (1990)

S.Takeda 等人：“培养的烟草细胞中积累的多肽的表征”Plant Cell Physiol.31。

N.Koizumi et al.: "Sequence analysis of cDNA encoding phosphoenolphruvate carboxylase from cultures tobacco cells" Plant Mol.Biology Plant M. 17. 535-593 (53)

N.Koizumi 等人：“对烟草细胞培养物中编码磷酸烯醇酸羧化酶的 cDNA 进行序列分析” Plant Mol.Biology Plant M. 17. 535-593 (53)

S.Takeda: "Characterization of polypeptides that accumulate in cultured Nicotiana tabacum cells" Plant & Cell Physiology. 31. 215-221 (1990)

S.Takeda：“培养的烟草细胞中积累的多肽的表征”植物

S. Takeda, F. Sato, K. Ida, Y. Yamada: "Characterization of polypeptides that accumulate in cultured Nicotiana tabacum cells" Plant Cell Physiol.31. 215-221 (1990)

S. Takeda、F. Sato、K. Ida、Y. Yamada：“在培养的烟草细胞中积累的多肽的表征”Plant Cell Physiol.31。

F. Sato, J. H. Fitchen, N, Takeshita, T. Hashimoto, N. Okada,: "Synthesis of Plant triosephosphate isomerase in Escherichia coli" Agric. Biol. Chem.54. 2189-2191 (1990)

F. Sato、J. H. Fitchen、N、Takeshita、T. Hashimoto、N. Okada，：“大肠杆菌中植物磷酸丙糖异构酶的合成”Agric。