Molecular basis of the adaptor domains from AAA-ATPases for locating organelles.

用于定位细胞器的 AAA-ATP 酶接头结构域的分子基础。

• 批准号: 16570136
• 负责人: HIROAKI Hidekazu
• 金额: $ 2.3万
• 依托单位: Yokohama City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16570136/
• 关键词: functional protein domains; Nuclear magnetic resonance (NMR); protein domain anatomy; late endosome; tubulin; cell skeleton; MIT domain; sequence identity; プロテアソーム; ポリユビキチン; ペルオキシソーム; X線結晶構造解析

AAA-ATPases are well conserved molecular machines from bacteria to human, which alters the conformations and/or multimerization states of substrate proteins with an ATP-depending manner. AAA-ATPases are characterized as harboring one or several "AAA" domains at their central region of the molecules, whereas various N-terminus domains may be responsible for differentiating their specific functions. I have hypothesized that some of them can be responsible not only for their substrate specificity, but also for their subcellular location. In order to solve this problem, so-called "protein domain anatomy" approach was taken.As a result, first we have identified a novel domain from the PEX1 N-terminal region. Surprisingly, although below 15% of sequence identity found to VCP and NSF, the structure of PEX1NTD is very similar to those corresponding domains. Second, we have identified the solution structures from N-terminal MIT domain of Vps4 type I AAA-ATPase and its distant homolog katanin p60. These domains adopted into an up-and-down three helix bundle fold with a characteristic cleft on the molecular surface, which may act as a ligand binding site, katanin p60 N-terminal domain was associated with a coiled-coil region, which should be designed prior to structural analysis. Some other AAA-ATPases require a shuttle factor molecule instead of its own adaptor domain. From one of them, we determined the solution structure of UBA domain from Dsk2 protein, an adaptor of 19S proteosome, in complex with ubiquitin. In addition, molecular rigidity and flexibility of K48- and K63-linked tetra ubiquitin chain were compared by using solution NMR.Finally, two domains of the five examined systems were found to have affinities to phosphoinositides. This suggested the domains share a function for associating organellar membranes.

AAA-ATP酶是从细菌到人的保守分子机器，它以降压方式改变了底物蛋白的构象和/或多聚化状态。 AAA-ATP酶的特征是在其分子的中央区域含有一个或几个“ AAA”域，而各种N末端域可能负责区分其特定功能。我假设其中一些不仅可以负责其底物特异性，而且还可以负责其亚细胞位置。为了解决这个问题，采用了所谓的“蛋白质结构域解剖”方法。结果，首先，我们从PEX1 N末端区域确定了一个新的域。令人惊讶的是，尽管在VCP和NSF中发现的序列身份的15％以下，但PEX1NTD的结构与那些相应的域非常相似。其次，我们从VPS4型AAA-ATPase的N末端MIT结构域及其远处的同源性Katanin P60确定了溶液结构。这些结构域在分子表面上具有特征性裂缝的上下三个螺旋束折叠，可以用作配体结合位点，katanin p60 n末端结构域与盘绕的螺旋线区域相关联，该区域应在结构分析之前设计。其他一些AAA-ATP酶需要穿梭因子分子而不是其自己的适配器域。从其中一个中，我们从DSK2蛋白（19S蛋白体的衔接子）中确定了UBA结构域的溶液结构，与泛素复杂。另外，通过使用溶液NMR比较了K48和K63连接的四泛素链的分子刚度和柔韧性，发现五个检查系统的两个结构域与磷酸肌醇具有亲和力。这表明这些域具有关联细胞器膜的功能。

项目成果

High-throughput construction method for expression vector of peptides for NMR study suited for isotopic labeling.

• DOI: 10.1093/protein/gzh044
• 发表时间: 2004-04
• 期刊: Protein engineering, design & selection : PEDS
• 作者: T. Tenno;Natsuko Goda;Y. Tateishi;H. Tochio;M. Mishima;H. Hayashi;M. Shirakawa;H. Hiroaki

ペルオキシソーム因子PEX1 ATPaseのN末端ドメインの立体構造が示したAAA ATPase間の進化的類縁関係

过氧化物酶体因子 PEX1 ATP 酶 N 端结构域的三维结构揭示了 AAA ATP 酶之间的进化关系

• 发表时间: 2006
• 期刊: 生物物理 46(3)
• 作者: 廣明 秀一;塩澤久美子;富井健太郎
• 通讯作者: 富井健太郎

A vector suited to high-throughput expression of isotopically-labeled peptides.

适合同位素标记肽高通量表达的载体。

• 发表时间: 2004
• 期刊: Protein Eng Design Sel. 17
• 作者: Tenno;T.;Goda;N.;Tateishi;Y.;Tochio;H.;Mishima;M.;Hayashi;H.;Shirakawa;M.;Hiroaki;H^*.
• 通讯作者: H^*.

Continuous Designed Cation-pi Interactions Increase the Thermal Stability of an α-Helical Coiled-coil.

连续设计的阳离子-π 相互作用提高了 α 螺旋线圈的热稳定性。

• 发表时间: 2005
• 期刊: Angewandte Chemie International Edition 44
• 作者: Sakurai;Y.;Mizuno;T.;Hiroaki;H.;Gohda;K.;Oku;J.I.;Tanaka;T^*.
• 通讯作者: T^*.

Optimization of size complementary of the aromatic side chain in the hydrophobic core of the designed coiled-coil.

优化所设计的卷曲线圈疏水核心中芳香族侧链的尺寸互补。

• 发表时间: 2005
• 期刊: J Pept Res 66
• 作者: Sakurai;Y.;Mizuno;T.;Hiroaki H.;Oku;J.I.;Tanaka;T^*
• 通讯作者: T^*