RAP AS A MOLECULAR CHAPERONE/ESCORT PROTEIN FOR LRP

LDL受体相关蛋白（LRP）是一个非常大的（600，000 kDa） 单链蛋白，具有四个相关的胞外结构域， 由多个（分别为2、8、10、11个）富含半胱氨酸的配体组成的转角 绑定重复。LRP已成为一种独特的内吞受体，由于其 能够结合和内吞许多结构和功能上 不同的配体。这些包括载脂蛋白E，2-巨球蛋白，tPA， 和尿激酶-纤溶酶原激活剂。一种39 kDa受体相关蛋白 (RAP)与细胞内LRP共纯化并阻断配体与LRP结合 体外本提案提供的初步数据表明， 不同的配体与LRP上的多个不同区域结合， RAP上的结合位点。此外，许多合成LRP和RAP的细胞 也可以合成受体的配体，这可以阻止正确的 LRP的折叠，如果它们与早期结合。私家侦探假设RAP 在LRP的折叠和正确运输中起着陪伴作用 通过结合它并防止与配体过早结合。 提出实验来阐明RAP的分子机制， 与LRP相互作用，通过解决其 晶体结构，确定RAP作为折叠分子伴侣的作用， 在体内和体外的方法，并确定运输的RAP和LRP 既作为单个分子又作为相互作用的复合物。

The LDL receptor-related protein (LRP) is an extremely large (600,000 kDa) single chain protein with four related extracellular domains that are in turn composed of multiple (2, 8, 10, 11 respectively) cysteine-rich ligand binding repeats. LRP has emerged as a unique endocytic receptor due to its ability to bind and endocytose a number of structurally and functionally distinct ligands. These include apolipoprotein E, 2-macroglobulin, tPA, and urokinase-plasminogen activator. A 39kDa receptor-associated protein (RAP) co-purifies with intracellular LRP and blocks ligand binding to LRP in vitro. Preliminary data provided in this proposal demonstrate that the different ligands bind to multiple and distinct regions on LRP and LRP binding sites on RAP. In addition, many cells that synthesize LRP and RAP also synthesize ligands for the receptor, which can prevent the correct folding of LRP if they associate to early. The P.I. hypothesizes that RAP serves a chaperoning function in the folding and proper trafficking of LRP by binding it and preventing premature association with ligand. Experiments are proposed to elucidate the molecular mechanism by which RAP interacts with LRP, explore the structural features of RAP by solving its crystal structure, determine the role of RAP as a folding chaperone using in vivo and in vitro methods, and define the trafficking of RAP and LRP both as individual molecules and interacting complexes.