Basu S K, Goldstein J L, Brown M S
J Biol Chem. 1978 Jun 10;253(11):3852-6.
An ultracentrifugation assay has been developed to measure low density lipoprotein (LDL) receptor activity in membranes prepared from cultured human fibroblasts. The binding site for 125I-labeled LDL in isolated membranes reflected the properties of the LDL receptor previously demonstrated in intact fibroblasts. It exhibited high affinity (Kd approximately 4 microgram of LDL protein/ml), specificity (LDL approximately 400-fold more effective than high density lipoprotein in competing with 125I-LDL for the binding site), dependence on calcium, and susceptibility to destruction by pronase. The number of LDL receptors detected in the in vitro membrane binding assay was similar to the number detected in intact cells. The number of receptors was reduced in membranes from fibroblasts that were grown in the presence of 25-hydroxycholesterol plus cholesterol and in fibroblast membranes from a subject with homozygous familial hypercholesterolemia, two situations in which the number of LDL receptors in intact fibroblasts is known to be reduced. The availability of a membrane binding assay that faithfully reflects the properties of the physiologic LDL receptor of intact cells should permit the characterization of this receptor in organs from intact humans and animals.
已开发出一种超速离心测定法，用于测量从培养的人成纤维细胞制备的膜中低密度脂蛋白（LDL）受体的活性。分离膜中125I标记的LDL的结合位点反映了先前在完整成纤维细胞中证明的LDL受体的特性。它表现出高亲和力（Kd约为4微克LDL蛋白/毫升）、特异性（LDL在与125I-LDL竞争结合位点方面比高密度脂蛋白有效约400倍）、对钙的依赖性以及对链霉蛋白酶破坏的敏感性。在体外膜结合测定中检测到的LDL受体数量与在完整细胞中检测到的数量相似。在25-羟胆固醇加胆固醇存在下生长的成纤维细胞的膜中以及来自纯合子家族性高胆固醇血症患者的成纤维细胞膜中，受体数量减少，已知在这两种情况下完整成纤维细胞中LDL受体的数量会减少。一种能如实反映完整细胞生理性LDL受体特性的膜结合测定法的可用性，应能使人们对来自完整人类和动物器官中的这种受体进行表征。
