Functional analysis of mammalian actin regulatory proteins, cofilin and destrin

哺乳动物肌动蛋白调节蛋白、丝切蛋白和结蛋白的功能分析

• 批准号: 02454534
• 负责人: NISHIDA Eisuke
• 金额: $ 3.84万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1992
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-02454534/
• 关键词: Actin-binding Proteins; Cytoskeleton; Phospholipase C; Inositol Phospholipid; Yeast; Dodecapeptide; アクチン結合蛋白質; アクチン結合部位; PIP_2; ホスホリパ-ゼC; イノシト-ルリン脂質; アクチン調節蛋白質; 化学架橋; 合成ペプチド

Cofilin and destrin, low molecular weight actin-binding proteins, are related to each other. Cofilin depolymerizes F-actin in a pH- dependent manner, while destrin does in a pH-independent manner. We have found that the Trp^<104>-Met^<115> region of cofilin (and destrin) is an actin binding site, and that this region is a polyphosphoinositide-binding site. The synthetic dodecapeptide patterned on the sequence corresponding to this region binds to PIP2 (PIP) as well as actin tightly, and thus is a PIP2 (PIP)-sensitive inhibitor of actin polymerization. We have further found that cofilin and the dodecapeptide inhibit PIP2 hydrolysis catalyzed by phospholipase C. Therefore, cofilin may function not only as a regulator of actin cytoskeleton but also as a regulator of the phosphoinositide signaling pathway.A cofilin-like protein has been identified from budding yeast. We carried out molecular cloning of this protein. We then expressed and purified a recombinant form of this protein, and characterized the recombinant protein. The results have shown that the protein depolymerizes F-actin in a pH-dependent manner, binds to both F- and G- actin, and binds to PIP2. These characteristics of the protein are completely the same as those of mammalian cofilin. Thus, we named this protein COF1. COF1 is found to be essential for yeast cell growth.

Cofilin和Destrin，低分子量肌动蛋白结合蛋白，彼此相关。 Cofilin以pH-依赖性的方式解聚了F-肌动蛋白，而Destrin则以pH不依赖的方式进行。我们发现，Cofilin（和Destrin）的TRP^<104> -MET^<115>是肌动蛋白结合位点，并且该区域是一个poly磷酸固醇结合位点。以对应于该区域的序列形成的合成十二肽与PIP2（PIP）以及肌动蛋白紧密结合，因此是肌动蛋白聚合的敏感抑制剂。我们进一步发现，磷脂酶C催化催化的Cofilin和十二肽抑制PIP2水解。因此，Cofilin不仅可以作为肌动蛋白细胞骨骼的调节剂起作用，而且还可以作为磷酸磷酸蛋白质信号途径的调节剂。萌芽的酵母。我们对该蛋白进行了分子克隆。然后，我们表达并纯化了该蛋白质的重组形式，并表征了重组蛋白。结果表明，蛋白质以pH值依赖性方式解聚，与F-和g-肌动蛋白结合并与PIP2结合。蛋白质的这些特征与哺乳动物cofilin的特征完全相同。因此，我们命名了该蛋白COF1。发现COF1对于酵母细胞生长至关重要。

项目成果

Yonezawa, N. et al.: "Inhibition of actin polymerization by a synthetic dodecapeptide patterned on the sequence around the actin- binding site of cofilin." J. Biol. Chem.266. 10485-10489 (1991)

Yonezawa, N. 等人：“通过合成十二肽抑制肌动蛋白聚合，该十二肽以肌丝蛋白丝切蛋白肌动蛋白结合位点周围的序列为图案。”

Yonezawa, N. et al.: "A short sequence responsible for both phosphoinositide binding and actin binding activities of cofilin." J. Biol. Chem.266. 17218-17221 (1991)

Yonezawa, N. 等人：“负责肌动蛋白丝切蛋白的磷酸肌醇结合和肌动蛋白结合活性的短序列。”

Moriyama, K. et al.: "Mutational analysis of an actin-binding site of cofilin and characterization of chimeric proteins between cofilin and destrin." J. Biol. Chem.267. 7240-7244 (1992)

Moriyama, K. 等人：“肌动蛋白丝切蛋白肌动蛋白结合位点的突变分析以及肌动蛋白丝切蛋白和结蛋白之间嵌合蛋白的表征。”

Moriyama,K.: "Mutational analysis of an actin-binding site of cofilin and characterization of chimeric proteins between cofilin and destrin" J.Biol.Chem.267. 7240-7244 (1992)

Moriyama,K.：“肌动蛋白丝切蛋白肌动蛋白结合位点的突变分析以及肌动蛋白丝切蛋白和结精蛋白之间嵌合蛋白的表征”J.Biol.Chem.267。

Yonezawa,N.: "Inhibition of the interactions of cofilin,destrin,and deoxyribonuclease I with actin by phosphoinositides" J.Biol.Chem.265. 8382-8386 (1990)

Yonezawa,N.：“磷酸肌醇抑制丝切蛋白、结蛋白和脱氧核糖核酸酶 I 与肌动蛋白的相互作用”J.Biol.Chem.265。