Molecular Recognition by Clathrin Adaptors

网格蛋白适配器的分子识别

• 批准号: 8553447
• 负责人: James Hurley
• 金额: $ 21.05万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8553447
• 关键词: Adaptor Signaling Protein; Affinity; Binding; Binding Sites; Biogenesis; Caliber; Clathrin; Clathrin Adaptors; Clathrin-Coated Vesicles; Complex; Endocytosis; Endosomes; Eukaryotic Cell; Goals; Individual; Lysosomes; Measures; Melanosomes; Nervous System Physiology; Organelles; Play; Property; Role; SNAPIN gene; Sorting - Cell Movement; Structure; Tubular formation; Vesicle; flexibility; golgi associated, gamma adaptin homologous, ADP-ribosylation factor interacting protein; molecular recognition; nervous system disorder; protein complex

Molecular Recognition by Clathrin Adaptors The clathrin coat plays a ubiquitous and fundamental role in endocytosis and in endosomal sorting within the eukaryotic cell. Clathrin forms a cage that surrounds cargo-bearing vesicles, but clathrin itself does not directly bind to cargo. Cargo is sorted into clathrin-coated vesicles by adaptor proteins that physically bridge cargo and clathrin. The best-known general purpose adaptors are the heterotetrameric adaptor protein complexes (AP complexes) and the multimodular GGA adaptor proteins. The overall goals of this project are 1) to identify the binding sites for cargo on adaptor proteins and measure their affinities quantitatively; 2) to determine the crystal structures of complexes between adaptors and soluble cargo fragments; and 3) to relate structure and function using mutational analysis. BLOC-1 (biogenesis of lysosome-related organelles complex-1) is critical for melanosome biogenesis and has also been implicated in neurological function and disease. BLOC-1 is not a clathrin adaptor, but functions in melanosome biogenesis in concert with the clathrin adaptor complex AP-3. We found that BLOC-1 is an elongated complex that contains one copy each of the eight subunits pallidin, Cappuccino, dysbindin, Snapin, Muted, BLOS1, BLOS2, and BLOS3. The complex appears as a linear chain of eight globular domains, ∼300 Å long and ∼30 Å in diameter. The individual domains are flexibly connected such that the linear chain undergoes bending by as much as 45. Two stable subcomplexes were defined, pallidin-Cappuccino-BLOS1 and dysbindin-Snapin-BLOS2. Both subcomplexes are 1:1:1 heterotrimers that form extended structures as indicated by their hydrodynamic properties. The two subcomplexes appear to constitute flexible units within the larger BLOC-1 chain, an arrangement conducive to simultaneous interactions with multiple BLOC-1 partners in the course of tubular endosome biogenesis and sorting.

笼状蛋白适配器的分子识别作用 在真核细胞的内吞作用和胞体分选中，网状蛋白被层起着普遍而基本的作用。网状蛋白形成一个笼子，包裹着载货的小泡，但网状蛋白本身并不直接与货物结合。货物通过连接货物和笼蛋白的接头蛋白被分类到覆盖着笼状蛋白的小泡中。最著名的通用接头是异四聚体接头蛋白复合体(AP复合体)和多模块GGA接头蛋白。该项目的总体目标是1)确定接头蛋白上货物的结合位点并定量测量它们的亲和力；2)确定接头与可溶性货物片段之间的复合体的晶体结构；3)利用突变分析将结构和功能联系起来。 BLOC-1(溶酶体相关细胞器复合体的生物发生)是黑素小体生物发生的关键，也与神经功能和疾病有关。BLOC-1不是一个网状蛋白适配器，但在黑素小体的生物发生中与网状蛋白适配器复合体AP-3协同发挥作用。我们发现BLOC-1是一个拉长的复合体，它包含Pallidin、Cappuccino、dybindin、Snapin、muted、BLOS1、BLOS2和BLOS3八个亚基中的一个拷贝。该复合体呈线性链状，由八个球状结构域组成，长300&8491；，直径30&8764；各个区域被灵活地连接，使得线性链经历多达45%的弯曲。定义了两个稳定的亚络合物：Pallidin-Cappuccino-BLOS1和dybindin-Snapin-BLOS2。这两个亚络合物都是1：1：1的杂三聚体，它们的流体力学性质表明它们形成了延伸的结构。这两个亚复合体似乎构成了更大的BLOC-1链中的灵活单元，这一安排有利于在管状内含体的生物发生和分选过程中与多个BLOC-1伙伴同时相互作用。