Mechanism of Coupling ATP hydrolysis to Ca^<2+> transport in sarcoplasmic reticulum Ca^<2+>-ATPase

肌浆网Ca^2-ATP酶中ATP水解与Ca^2转运的耦合机制

• 批准号: 14580619
• 负责人: DAIHO Takashi
• 金额: $ 2.37万
• 依托单位: Asahikawa Medical College
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14580619/
• 关键词: sarcoplasmic reticulum; Ca^<2+>-ATPase; phosphorylated intermediate; Daner's disease; Active transport; SERCA; calcium pump; P-type ATPase; P-type ATPase; ATPase; ドメイン間相互作用; 界面活性剤; 削除変異; loop

1.Mg/F/E(Enzyme) complex of sarcoplasmic reticulum Ca^<2+>-ATPase is an stable analog of phosphorylated intermediate(EP). We have found that the solubilized and delipidated Mg/F/E is remarkably stable against denaturation. 2.The Ca^<2+>-ATPase has 3 cytoplasmic domains (A,P,N). We have found that interaction between Val^<200> loop of A domain and P domain is important for Ca^<2+>-translocation from EP.3.Glu40-Ser48 loop connect A domain and the 1st transmembrane segment. We have found that the adequate length of this loop is important for conformational transition of EP, which is critical for Ca^<2+> translocation. We found that some mutations in this loop region reported in the Darier's disease pedigrees will also cause inhibition of EP transition. 4.We have found that Arg^<334> is important for EP transition and Tyr^<122> for Ca^<2+> translocation after the transition of EP. We have proposed a model for coupling, in which energy of ATP hydrolysis is transmitted to Ca^<2+> transport site through the large movements of cytoplasmic domains. 5.We have shown that the complexes Be/F/E,Al/F/E, and Mg/F/E are stable analogs for the ground state, transition state, and enzyme/product in EP hydrolysis. We also found that the structural changes in the catalytic site in EP hydrolysis will close the lumenal gate for Ca^<2+> translocation to prevent Ca^<2+> leak form the lumen.

1.肌浆网Ca^（2+）-ATP酶的Mg/F/E（酶）复合物是磷酸化中间体（EP）的稳定类似物。我们已经发现，增溶和脱脂的Mg/F/E对变性非常稳定。2. Ca^<2+>-ATP酶有3个胞质结构域（A、P、N）。我们发现<200>A结构域的瓦尔环与P结构域的相互作用对Ca^&lt;2+&gt;-转运起重要作用。我们已经发现，该环的足够长度对于EP的构象转变是重要的，而EP的构象转变对于Ca^&lt;2+&gt;转运是至关重要的。我们发现在Darier病家系中报道的这个环区的一些突变也会导致EP转换的抑制。4.发现Arg^<334>在EP转换中起重要作用，Tyr^<122>在EP转换后Ca^2+转运中起重要作用。我们提出了一个耦合模型，其中ATP水解的能量通过胞质结构域的大运动传递到Ca^2+转运位点。5.我们证明了Be/F/E、Al/F/E和Mg/F/E配合物是EP水解的基态、过渡态和酶/产物的稳定类似物。我们还发现EP水解过程中催化位点的结构变化将关闭Ca^2+转运的腔门，阻止Ca^2+从腔中漏出。

项目成果

Takashi Daiho, Kazuo Yamasaki, Guoli Wang, Stefania Danko, Hajime Iizuka, Hiroshi Suzuki: "Deletions of Any Single Residues in Glu^<40>-Ser^<48> Loop Connecting A Domain and the First Transmembrane Helix of Sarcoplasmic Reticulum Ca^<2+>-ATPase Result in

Takashi Daiho、Kazuo Yamasaki、Guoli Wang、Stefania Danko、Hajime Iizuka、Hiroshi Suzuki：“连接结构域和肌浆网 Ca^ 的第一个跨膜螺旋的 Glu^<40>-Ser^<48> 环中任何单个残基的删除

Kazuo Yamasaki, Takashi Daiho, Stefania Danko, Hiroshi Suzuki: "Multiple and distinct effects of mutations of Tyr^<122>, Glu^<123>, Arg^<329>, and Arg^<334> involved in interactions between the top part of second and fourth transmembrane helices in Sarcop

Kazuo Yamasaki、Takashi Daiho、Stefania Danko、Hiroshi Suzuki：“Tyr^<122>、Glu^<123>、Arg^<329> 和 Arg^<334> 突变的多重且独特的影响涉及顶部之间的相互作用

Stefania Danko, Kazuo Yamasaki, Takashi Daiho, Hiroshi Suzuki: "Distinct Natures of Be/F-bound, Al/F-bound, and Mg/F-bound Stable Analogues of ADP-insensitive Phosphoenzyme Intermediate of Sarcoplasmic Reticulum Ca^<2+>-ATPase : Changes in catalytic and t

Stefania Danko、Kazuo Yamasaki、Takashi Daiho、Hiroshi Suzuki：“肌浆网 Ca^2 的 ADP 不敏感磷酸酶中间体的 Be/F 结合、Al/F 结合和 Mg/F 结合稳定类似物的独特性质

Kazuo Yamasaki, Takashi Daiho, Hiroshi Suzuki: "Remarkable stability of solubilized and delipidated sarcoplasmic reticulum Ca^<2+>-ATPase with tightly bound fluoride and magnesium against detergent induced denaturation"The Journal of Biological Chemistry.

Kazuo Yamasaki、Takashi Daiho、Hiroshi Suzuki：“溶解和脱脂的肌浆网 Ca^2-ATP 酶与紧密结合的氟化物和镁对洗涤剂诱导的变性具有显着的稳定性”《生物化学杂志》。

山崎 和生, 大保 貴嗣, 鈴木 裕: "筋小胞体Ca^<2+>-ATPaseの細胞質ドメインに存在するイオンペアの役割"生化学. 74・8. 1023 (2002)

Kazuo Yamazaki、Takashi Oho、Yutaka Suzuki：“肌浆网 Ca^<2+>-ATP 酶的细胞质结构域中存在的离子对的作用”生物化学 74・8。