Transfer of overexpressed protein into organelle and its regulation mechanism

过表达蛋白转入细胞器及其调控机制

• 批准号: 04680157
• 负责人: AOYAMA Toshifumi
• 金额: $ 1.28万
• 依托单位: Shinshu 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: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04680157/
• 关键词: acyl-CoA dehydrogenase; acyl-CoA oxidase; PMP70; vaccinia virus; overexpression; stability in organelle; cDNA-expression; peroxisome; ワクシニアウイルスベクター; ペルオキシソームへの移行; Zellweger細胞; 極長鎖脂肪酸アシルCoA脱水素酵素; mRNAの安定性

cDNA encoding mitochondrial very-long-chain acyl-CoA dehydrogenase was cloned. Amino acid sequence indicated that the enzyme had about 180 amino acids of extra polypeptide at the carboxyl terminal side, when it was compared with the other acyl-CoA dehydrogenases. A series of mutants, deleting about 60 amino acids of polypeptide from carboxyl terminus, were inserted into vaccinia virus to construct recombinant viruses. Expressed protein in rat hepatoma H4 was examined by means of immunoblot analysis. Mutant protein was less stable in mitochondria than mature protein concerning period after the expression. Mutant protein also lost the features of membrane binding and association to form dimer. These results indicate that the extra polypeptide at carboxyl side has unique function to stabilize the protein in mitochondria. Similar results are obtained concerning the extra polypeptide at carboxyl side of peroxisomal acyl-CoA oxidase.

克隆了编码线粒体极长链酰基辅酶A脱氢酶的cDNA。氨基酸序列表明，与其他酰基辅酶A脱氢酶相比，该酶在羧基端有约180个氨基酸的额外多肽。将一系列从羧基末端删除约60个氨基酸的多肽突变体插入牛痘病毒中构建重组病毒。通过免疫印迹分析检查大鼠肝癌H4中表达的蛋白。就表达后的一段时间而言，突变蛋白在线粒体中的稳定性低于成熟蛋白。突变蛋白也失去了膜结合和缔合形成二聚体的特征。这些结果表明羧基侧额外的多肽具有稳定线粒体中蛋白质的独特功能。关于过氧化物酶体酰基辅酶A氧化酶羧基侧的额外多肽，获得了类似的结果。

项目成果

Miwako Ozaki, Kiyoshi Matsumura, Shuji Kaneko, Masamichi Sato, Yasuyoshi Watanabe, and Toshifumi Aoyama: "A vaccinia virus vector for efficiently introducing into hippocampal slices" Biochem.Biophys.Res.Commun.193 (1993)

Miwako Ozaki、Kiyoshi Matsumura、Shuji Kaneko、Masamichi Sato、Yasuyoshi Watanabe 和 Toshifumi Aoyama：“一种用于有效导入海马切片的痘苗病毒载体”Biochem.Biophys.Res.Commun.193 (1993)

Takehiko Kamijo, Ronald J.A.Wanders, Jean-Marie Saudubray, Toshifumi Aoyama, Atsushi Komiyama, and Takashi Hashimoto: "Biochemical and molecular heterogeneity of mutant trifunctional protein in fibroblasts from two patients with long-chain 3-hydroxyacyl-C

Takehiko Kamijo、Ronald J.A.Wanders、Jean-Marie Saudubray、Toshifumi Aoyama、Atsushi Komiyama 和 Takashi Hashimoto：“两名长链 3-羟基酰基-C 患者的成纤维细胞中突变三功能蛋白的生化和分子异质性

Toshifumi Aoyama,Yasushi Uchida,Michael Marble,Karen Hofman,William,J.Rhead,and Takashi Hashimoto: "A novel discase with deficiency of mitochondrial very-long-chain acyl-CoA dehydrogenase" Biochem.Biophys.Res.Commun.191. 1369-1372 (1993)

Toshifumi Aoyama、Yasushi Uchida、Michael Marble、Karen Hofman、William、J.Rhead 和 Takashi Hashimoto：“线粒体极长链酰基辅酶 A 脱氢酶缺乏的新型疾病”Biochem.Biophys.Res.Commun.191。

Toshifumi Aoyama,Yasushi Uchida,Michael Marble,Karen Hofman,William,J.Rhead,and Takashi Hashimoto: "A novel disease with deficiency of mitochondrial very-long-chain acyl-CoA dehydrogenase" Biochem.Biophys.Res.Commun.191. 1369-1372 (1993)

Toshifumi Aoyama、Yasushi Uchida、Michael Marble、Karen Hofman、William、J.Rhead 和 Takashi Hashimoto：“线粒体极长链酰基辅酶 A 脱氢酶缺乏的新型疾病”Biochem.Biophys.Res.Commun.191。

Toshifumi Aoyama, Ichiro Ueno, Takehiko Kamijo, and Takashi Hashimoto: "Rat very-long-chain acyl-CoA dehydrogenase, a novel mitochondrial acyl-CoA dehyrogenase gene product, is a rate-limiting enzyme in long-chain fatty acid beta-oxidationsystem : cDNA an

Toshifumi Aoyama、Ichiro Ueno、Takehiko Kamijo 和 Takashi Hashimoto：“大鼠极长链酰基辅酶A脱氢酶是一种新型线粒体酰基辅酶A脱氢酶基因产物，是长链脂肪酸β-氧化系统中的限速酶