Medh J D, Fry G L, Bowen S L, Pladet M W, Strickland D K, Chappell D A
Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242.
J Biol Chem. 1995 Jan 13;270(2):536-40. doi: 10.1074/jbc.270.2.536.
The 39-kDa receptor-associated protein (RAP) is cosynthesized and co-purifies with the low density lipoprotein receptor-related protein (LRP)/alpha 2-macroglobulin receptor and is thought to modulate ligand binding to LRP. In addition to binding LRP, RAP binds two other members of the low density lipoprotein (LDL) receptor family, gp330 and very low density lipoprotein (VLDL) receptors. Here, we show that RAP binds to LDL receptors as well. In normal human foreskin fibroblasts, RAP inhibited LDL receptor-mediated binding and catabolism of LDL and VLDL with Sf 20-60 or 100-400. RAP inhibited 125I-labeled LDL and Sf 100-400 lipoprotein binding at 4 degrees C with KI values of 60 and 45 nM, respectively. The effective concentrations for 50% inhibition (EC50) of cellular degradation of 2.0 nM 125I-labeled LDL, 4.7 nM 125I-labeled Sf 20-60, and 3.6 nM 125I-labeled Sf 100-400 particles were 40, 70, and 51 nM, respectively. Treatment of cells with lovastatin to induce LDL receptors increased cellular binding, internalization, and degradation of RAP by 2.3-, 1.7-, and 2.6-fold, respectively. In solid-phase assays, RAP bound to partially purified LDL receptors in a dose-dependent manner. The dissociation constant (KD) of RAP binding to LDL receptors in the solid-phase assay was 250 nM, which is higher than that for LRP, gp330, or VLDL receptors in similar assays by a factor of 14 to 350. Also, RAP inhibited 125I-labeled LDL and Sf 100-400 VLDL binding to LDL receptors in solid-phase assays with KI values of 140 and 130 nM, respectively. Because LDL bind via apolipoprotein (apo) B100 whereas VLDL bind via apoE, our results show that RAP inhibits LDL receptor interactions with both apoB100 and apoE. These studies establish that RAP is capable of binding to LDL receptors and modulating cellular catabolism of LDL and VLDL by this pathway.
39千道尔顿的受体相关蛋白（RAP）与低密度脂蛋白受体相关蛋白（LRP）/α2-巨球蛋白受体共同合成并共同纯化，被认为可调节配体与LRP的结合。除了结合LRP外，RAP还结合低密度脂蛋白（LDL）受体家族的另外两个成员，即gp330和极低密度脂蛋白（VLDL）受体。在此，我们表明RAP也能结合LDL受体。在正常人包皮成纤维细胞中，RAP以Sf 20 - 60或100 - 400抑制LDL受体介导的LDL和VLDL的结合及分解代谢。RAP在4℃时抑制125I标记的LDL和Sf 100 - 400脂蛋白的结合，其抑制常数（KI）值分别为60和45 nM。对于2.0 nM 125I标记的LDL、4.7 nM 125I标记的Sf 20 - 60以及3.6 nM 125I标记的Sf 100 - 400颗粒，细胞降解的50%抑制有效浓度（EC50）分别为40、70和51 nM。用洛伐他汀处理细胞以诱导LDL受体，可使RAP的细胞结合、内化和降解分别增加2.3倍、1.7倍和2.6倍。在固相分析中，RAP以剂量依赖方式结合部分纯化的LDL受体。在固相分析中，RAP与LDL受体结合的解离常数（KD）为250 nM，这比在类似分析中与LRP、gp330或VLDL受体的解离常数高14至350倍。此外，在固相分析中，RAP抑制125I标记的LDL和Sf 100 - 400 VLDL与LDL受体的结合，其KI值分别为140和130 nM。由于LDL通过载脂蛋白（apo）B100结合，而VLDL通过apoE结合，我们的结果表明RAP抑制LDL受体与apoB100和apoE的相互作用。这些研究证实RAP能够结合LDL受体并通过该途径调节LDL和VLDL的细胞分解代谢。
