Vash B, Phung N, Zein S, DeCamp D
Departmnt of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX, USA.
Blood. 1998 Nov 1;92(9):3277-85.
The low-density lipoprotein receptor-related protein (LRP) is a 600-kD scavenger receptor that binds a number of protein ligands with high affinity. Although some ligands do not compete with each other, binding of all is uniformly blocked by the 39-kD receptor-associated protein (RAP). RAP is normally found in the endoplasmic reticulum and seems to function as a chaperone for LRP. To identify the binding sites for RAP, lactoferrin, and plasminogen activator inhibitor-1 (PAI-1), a bacterial expression system has been developed to produce soluble LRP fragments spanning residues 783-1399. These residues overlap most of the CNBr fragment containing the second cluster of complement-type repeats (C). Solid phase binding assays show that 125I-RAP binds to fragments containing three successive complement-type repeats: C5-C7. PAI-1 and lactoferrin bind to the same fragments. A fragment containing C5-C7 also blocks uptake and degradation of 125I-RAP by fibroblasts in a concentration-dependent manner. Binding competition experiments show that RAP, PAI-1, and lactoferrin each inhibit the binding of the others, suggesting that at this site in LRP, RAP acts as a competitive, rather than an allosteric, inhibitor of PAI-1 and lactoferrin binding.
低密度脂蛋白受体相关蛋白（LRP）是一种600kD的清道夫受体，能以高亲和力结合多种蛋白质配体。尽管一些配体之间不存在相互竞争，但39kD的受体相关蛋白（RAP）能统一阻断所有配体的结合。RAP通常存在于内质网中，似乎起着LRP伴侣蛋白的作用。为了确定RAP、乳铁蛋白和纤溶酶原激活物抑制剂-1（PAI-1）的结合位点，已开发出一种细菌表达系统，用于产生跨越783 - 1399位残基的可溶性LRP片段。这些残基覆盖了包含第二个补体样重复序列簇（C）的大部分溴化氰片段。固相结合试验表明，125I-RAP与包含三个连续补体样重复序列的片段：C5 - C7结合。PAI-1和乳铁蛋白与相同的片段结合。包含C5 - C7的片段也以浓度依赖的方式阻断成纤维细胞对125I-RAP的摄取和降解。结合竞争实验表明，RAP、PAI-1和乳铁蛋白各自抑制其他两者的结合，这表明在LRP的这个位点，RAP作为PAI-1和乳铁蛋白结合的竞争性抑制剂，而非变构抑制剂发挥作用。
