cDNAs and Antibodies for Small GTP-binding Proteins

小 GTP 结合蛋白的 cDNA 和抗体

• 批准号: 01870017
• 负责人: TAKAI Yoshimi
• 金额: $ 7.23万
• 依托单位: Kobe University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B).
• 财政年份: 1989
• 资助国家: 日本
• 起止时间: 1989 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01870017/
• 关键词: ras p21; small GTP-binding proteins; prenylation; GDP; GTP exchange protein; GTPase activating protein; Tumor markers; <ras>___ー p21; GTP結合蛋白質; smgp25; smgp21; GDI

There is a superfamily of ras p21/ras p21-like small GTP-binding proteins (small G proteins) with GTPase activity. Over forty members have been reported, and all of them with Mrs between 20,000 and 41,000 are composed of a single respective polypeptide. Evidence is accumulating that small G proteins are involved in the regulation of cell growth and differentiation. In this research project, we have investigated the C-terminal structures of small G proteins, purified the regulatory proteins for small G proteins, and made monoclonal antibodies against small G proteins and their regulatory proteins. Most of small G proteins have the unique consensus C-terminal motifs containing at least one cysteine residue. We have found that the C-terminal systeine residues of smg p21B, rhoA p21, and smg p25A are geranylgeranylated, and that these prenylation are essential for each small G protein to bind to membranes. We have partially purified and characterized geranyl-geranyl transferase for rhoA p21 … More from bovine brain. Small G proteins have two interconvertible forms, GDP-bound inactive and GTP-bound active forms. The conversion from the GDP-bound to GTP-bound form and the reverse conversion are induced by GDP/GTP exchange and GTPase reactions, respectively. We have purified and characterized several GDP/GTP exchange proteins (GDP dissociation stimulator (GDS) and GDP dissociation inhibitor (GDI)) and GTPase activating Proteins (GAP) for small G proteins. Among these regulatory proteins, we have cloned the cDNAs of smg p21 GDS, smg p25A GDI, and rho GDI, and made monoclonal antibodies against them. We have found that the levels of smg p21 mRNA increase in hematopoietic leukemiae cells after their differentiation into hematopoietic cells. We have also found that smg p25A is mainly detected inneural tissues by use of its specific monoclonal antibody, and that the levels of smg p25A mRNA increase in PC12 cells after their differentiation into neuron-like cells. These results suggest that smg p21 and smg p25A can be used as potential tumor markers for tumors derived from hematopoietic and neural tissues, respectively. Key Words : ras p21, small G proteins, Prenylation, GDP/GTP exchange protein, GTPase activating protein, Tumor markers. Less

具有GTPase活性的RAS P21/RAS样小GTP结合蛋白（小G蛋白）的超家族。据报道，已有40多名成员，所有成员的MRS在20,000至41,000之间都是由单个特异性多肽组成的。有证据表明，小的G蛋白参与细胞生长和分化的调节。在该研究项目中，我们研究了小G蛋白的C末端结构，纯化了小G蛋白的调节蛋白，并制造了针对小G蛋白及其调节蛋白的单克隆抗体。大多数小的G蛋白具有至少一个半胱氨酸保留率的独特共识C末端基序。我们发现，SMG P21B，RhoA P21和SMG P25A的C末端收缩蛋白保留是闪烷基凝集苯基的，并且这些原始化对于每个小G蛋白与膜结合至关重要。我们已经部分纯化并表征了rhoa p21的香烷基 - 凝视酶转移酶，其中来自牛脑。小的G蛋白具有两种可互通的形式，GDP结合和GTP结合的活性形式。 GDP/GTP交换和GTPase反应分别诱导了从GDP结合到GTP结合形式和反向转换的转换。我们已经纯化并表征了几种GDP/GTP交换蛋白（GDP解离刺激剂（GDS）和GDP解离抑制剂（GDI））和GTPase激活蛋白（GAP）的小G蛋白。在这些调节蛋白中，我们将SMG P21 GDS，SMG P25A GDI和Rho GDI的CDNA克隆，并对它们制成了单克隆抗体。我们发现，造血性白血病细胞中SMG P21 mRNA的水平在分化为造血细胞后的水平。我们还发现，SMG P25a主要通过使用其特异性单克隆抗体检测到内部组织，并且SMG P25a mRNA的水平在PC12细胞中分化为神经元样细胞后的水平增加。这些结果表明，SMG P21和SMG P25A可以用作分别源自造血和神经元组织的肿瘤的潜在肿瘤标记。关键词：RAS P21，小G蛋白，原始化，GDP/GTP交换蛋白，GTPase激活蛋白，肿瘤标记。较少的

项目成果

Sasaki,T.: "A mammalian inhibitory GDP/GTP exchange protein (GDI) for smg p25A is active on the yeast SEC4 protein." Mol.Cell.Biol.(1991)

Sasaki,T.：“smg p25A 的哺乳动物抑制性 GDP/GTP 交换蛋白 (GDI) 对酵母 SEC4 蛋白具有活性。”

Hata,Y.: "Inhibition of the ras p21 GTPaseーactivating proteinーstimulated GTPase activity of cーHaーras p21 by smg p21 having the same putative effector domain as ras p21s." J.Biol.Chem.265. 7104-7107 (1990)

Hata, Y.：“smg p21 与 ras p21s 具有相同的推定效应结构域，对 ras p21 GTPase 激活蛋白刺激的 c-Haras p21 GTPase 活性的抑制。J.Biol.Chem.265。” 1990）

Araki,S.: "Regulation of reversible binding of smg p25A,a ras p21ーlike GTPーbinding protein,to synaptic plasma membranes and vesicles by its specific regulatory protein,GDP dissociation inhibitor." J.Biol.Chem.265. 13007-13015 (1990)

Araki, S.：“smg p25A（一种 ras p21 样 GTP 结合蛋白）通过其特异性调节蛋白 GDP 解离抑制剂对突触质膜和囊泡的可逆结合进行调节。”J.Biol.Chem.265。 -13015 (1990)

Fukumoto, Y.: "Activation of the c-fos Serum-Response Element by the Activated c-Ha-ras Protein in a Manner Independent of Protein Kinase C and cAMP-Dependent Protein Kinase." J. Biol. Chem.265. 774-780 (1990)

Fukumoto, Y.：“激活的 c-Ha-ras 蛋白以独立于蛋白激酶 C 和 cAMP 依赖性蛋白激酶的方式激活 c-fos 血清反应元件。”

Yamamoto,J.: "A GTPaseーactivating protein for rhoB p20,a ras p21ーlike GTPーbinding proteinーpartial purification,characterization and subcellular distribution in rat brain." Mol.Brain Res.8. 105-111 (1990)

Yamamoto, J.：“rhoB p20 的 GTP 激活蛋白，一种 ras p21 样 GTP 结合蛋白 - 大鼠脑中的部分纯化、表征和亚细胞分布。”Mol.Brain Res.8（1990）。