Molecular Recognition by Clathrin Adaptors

网格蛋白适配器的分子识别

• 批准号: 8349738
• 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/8349738
• 关键词: Adaptor Protein Complex Subunits; Adaptor Signaling Protein; Affinity; Alzheimer' s Disease; Amino Acid Sequence; Amyloid beta-Protein; Amyloid beta-Protein Precursor; Binding; Binding Sites; Biochemical; Clathrin; Clathrin Adaptors; Clathrin-Coated Vesicles; Complex; Endocytosis; Endosomes; Eukaryotic Cell; Family member; Goals; Golgi Apparatus; Location; Measures; Mediating; Peptide Sequence Determination; Play; Process; Property; Proteins; Reporting; Roentgen Rays; Role; Signal Transduction; Sorting - Cell Movement; Structure; Tail; Vesicle; gamma secretase; golgi associated, gamma adaptin homologous, ADP-ribosylation factor interacting protein; molecular recognition; polarized cell; protein complex; trans-Golgi Network

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. Adaptor protein 4 (AP-4) is the most recently discovered and least well-characterized member of the family of heterotetrameric adaptor protein (AP) complexes that mediate sorting of transmembrane cargo in post-Golgi compartments. In the previous FY, we reported the interaction of an YKFFE sequence from the cytosolic tail of the Alzheimer's disease amyloid precursor protein (APP) with the mu4 subunit of AP-4. Biochemical and X-ray crystallographic analyses reveal that the properties of the APP sequence and the location of the binding site on mu4 are distinct from those of other signal-adaptor interactions. Disruption of the APP-AP-4 interaction decreases localization of APP to endosomes and enhances gamma-secretase-catalyzed cleavage of APP to the pathogenic amyloid-beta peptide. These findings demonstrate that APP and AP-4 engage in a distinct type of signal-adaptor interaction that mediates transport of APP from the trans-Golgi network (TGN) to endosomes, thereby reducing amyloidogenic processing of the protein. In this FY we focused on unique recognition mechanisms of AP complexes in polarized cells.

网格蛋白衔接子的分子识别 网格蛋白外壳在真核细胞内的内吞作用和内体分选中起着普遍存在的和基本的作用。网格蛋白形成一个笼，围绕货物轴承囊泡，但网格蛋白本身不直接结合货物。货物通过衔接蛋白被分选到网格蛋白包被的囊泡中，衔接蛋白物理桥接货物和网格蛋白。最著名的通用衔接子是异四聚体衔接蛋白复合物（AP复合物）和多模块GGA衔接蛋白。该项目的总体目标是：1）鉴定衔接蛋白上货物的结合位点并定量测量其亲和力; 2）确定衔接蛋白和可溶性货物片段之间复合物的晶体结构; 3）使用突变分析将结构和功能联系起来。 衔接蛋白4（AP-4）是最近发现的并且最不充分表征的异源四聚体衔接蛋白（AP）复合物家族的成员，其介导高尔基体后区室中跨膜货物的分选。在上一个财年，我们报告了从阿尔茨海默氏病淀粉样前体蛋白（APP）的胞质尾部的YKFFE序列与AP-4的mu 4亚基的相互作用。生化和X射线晶体学分析表明，APP序列的属性和穆4上的结合位点的位置是不同的其他信号适配器相互作用。APP-AP-4相互作用的破坏减少了APP在内体的定位，并增强了APP向致病性淀粉样蛋白β肽的γ-分泌酶催化的裂解。这些发现表明，APP和AP-4参与了一种独特类型的信号-适配器相互作用，介导APP从trans-Golgi网络（TGN）转运到内体，从而减少蛋白质的淀粉样蛋白加工。在本财年中，我们专注于极化细胞中AP复合物的独特识别机制。