The use of monoclonal antibodies to localize the low density lipoprotein receptor-binding domain of apolipoprotein B.
The use of monoclonal antibodies to localize the low density lipoprotein receptor-binding domain of apolipoprotein B.
复制标题
使用单克隆抗体定位载脂蛋白 B 的低密度脂蛋白受体结合域。
DOI：
10.1016/s0021-9258(19)47176-7
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发表时间：
1989
期刊：
影响因子：
--
通讯作者：
Y. Marcel
中科院分区：
文献类型：
--
作者：
R. Milne;R. Theolis;R. Maurice;R. Pease;P. Weech;E. Rassart;J. Fruchart;James F. Scott;Y. Marcel
人载脂蛋白（apo）B-100由4536个氨基酸组成。认为apoB与低密度脂蛋白（LDL）受体的结合涉及配体的碱性氨基酸与受体的酸性残基之间的相互作用。已经提出了三种替代模型来描述这种相互作用：1）apoB的单个区域参与受体结合; 2）来自apoB一级结构的碱性氨基酸组在apoB受体结合中协同作用;和3）apoB包含多个独立的结合区域。我们已经发现，特异性针对跨越apoB残基3249（T2/T3连接）处的凝血酶切割位点的区域的单克隆抗体（Mab）完全阻断LDL与LDL受体的结合。对该区域以外的表位具有特异性的单克隆抗体没有或部分阻断LDL结合的能力。为了确定直接参与与LDL受体相互作用的apoB区域，我们测试了22种不同的单克隆抗体与已经固定在受体上的LDL结合的能力。对位于残基2835和2922之间的表位具有特异性的单克隆抗体可以与固定在其受体上的LDL上的表位结合，而残基2980和3084之间的第二表位在受体结合的LDL上是不可接近的。apoB残基3500附近的一系列表位是完全不可接近的，另一个位于残基4027和4081之间的表位在受体结合的LDL上是难以接近的。相反，位于残基4154和4189之间的表位完全暴露。对区域3000-4000的上游和下游的表位具有特异性的单克隆抗体可以以接近1的化学计量结合受体结合的LDL。我们的研究结果强烈表明，apoB直接参与LDL-受体相互作用的独特区域是T2/T3连接。
Human apolipoprotein (apo) B-100 is composed of 4536 amino acids. It is thought that the binding of apoB to the low density lipoprotein (LDL) receptor involves an interaction between basic amino acids of the ligand and acidic residues of the receptor. Three alternative models have been proposed to describe this interaction: 1) a single region of apoB is involved in receptor binding; 2) groups of basic amino acids from throughout the apoB primary structure act in concert in apoB receptor binding; and 3) apoB contains multiple independent binding regions. We have found that monoclonal antibodies (Mabs) specific for a region that spans a thrombin cleavage site at apoB residue 3249 (T2/T3 junction) totally blocked LDL binding to the LDL receptor. Mabs specific for epitopes outside this region had either no or partial ability to block LDL binding. In order to define the region of apoB directly involved in the interaction with the LDL receptor we have tested 22 different Mabs for their ability to bind to LDL already fixed to the receptor. A Mab specific for an epitope situated between residues 2835 and 2922 could bind to its epitope on LDL fixed to its receptor whereas a second epitope between residues 2980 and 3084 is inaccessible on receptor-bound LDL. A series of epitopes near residue 3500 of apoB is totally inaccessible, and another situated between residues 4027 and 4081 is poorly accessible on receptor-bound LDL. In contrast, an epitope that is situated between residues 4154 and 4189 is fully exposed. Mabs specific for epitopes upstream and downstream of the region 3000-4000 can bind to receptor-bound LDL with a stoichiometry close to unity. Our results strongly suggest that the unique region of apoB directly involved in the LDL-receptor interaction is that of the T2/T3 junction.