Role of amphipathic transmembrane domains of single-spanning membrane proteins as determinants of their oligomeric state and subcellular localization.

单跨膜蛋白的两亲性跨膜结构域作为其寡聚状态和亚细胞定位的决定因素的作用。

• 批准号: 258126872
• 负责人: Dr. Andreas Ernst
• 金额: --
• 依托单位: Department of Cell Biology
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2014
• 资助国家: 德国
• 起止时间: 2013-12-31 至 2015-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/258126872?language=en
• 关键词: Role; amphipathic; transmembrane; domains; single

Molecular dynamics simulations hinted at an unusual polar interface in mediating TMD-TMD interactions for p24 proteins (Contreras-Ernst et al., 2012). Other reports hint at the involvement of similar polar faces of TMDs in mediating localization to the Golgi and protein-protein interactions (Machamer et al., 1993; Dawson et al., 2002, 2003). Preliminary data that arose from bioinformatical analysis suggests that amphipathic TMD design is common for single-spanning membrane proteins and enriched in the early secretory pathway. With respect to the different models that attempt to explain specific localization of enzymes to the Golgi, I am planning to investigate whether the amphipathicity of TMDs is a crucial determinant in achieving this function. This hypothesis might combine all existing models, as ER-localization would be achieved via cytoplasmic retrieval signals of ER resident membrane proteins, while plasma membrane residents that also possess amphipathic TMDs would, due to their increased hydrophobic length, be excluded from Golgienzyme oligomers (TMD-partitioning model). Within the Golgi, kin recognition of Golgi enzymes would then be achieved by TMD-based interactions among oligomers. To study the contribution of amphipathic TMDs in homo- and heterooligomerization, a FRET-based proteoliposomal system and an in vivo BIrA-based biotinylation assay will be employed. The role of the amphipathic TMD of the respective candidate in its subcellular localization will be addressed by two-color, high-resolution STED nanoscopy.

分子动力学模拟表明，在介导p24蛋白的TMD-TMD相互作用中存在不寻常的极性界面（Contreras-Ernst et al., 2012）。其他报告暗示，类似的tmd极性面参与介导高尔基体定位和蛋白-蛋白相互作用（Machamer等，1993；Dawson等，2002,2003）。来自生物信息学分析的初步数据表明，单跨膜蛋白的两型TMD设计是常见的，并且在早期分泌途径中丰富。对于试图解释酶在高尔基体中的特定定位的不同模型，我计划研究tmd的两致病性是否是实现这一功能的关键决定因素。这一假设可能结合所有现有的模型，因为内质网定位将通过内质网驻留膜蛋白的细胞质检索信号来实现，而同样具有两亲性tmd的质膜驻留蛋白由于其疏水长度增加，将被排除在高尔基酶低聚物（tmd分配模型）之外。在高尔基体中，高尔基酶的亲缘识别将通过低聚物之间基于tmd的相互作用来实现。为了研究两亲性tmd在同源和异源寡聚化中的作用，将采用基于fret的蛋白脂质体系统和基于bira的体内生物素化测定。两亲性TMD在其亚细胞定位中的作用将通过双色、高分辨率STED纳米显微镜来解决。