Allan C M, Tetaz T, Fidge N H
Protein Chemistry and Molecular Biology Unit, Baker Medical Research Institute, Prahran, Victoria, Australia.
J Lipid Res. 1991 Apr;32(4):595-601.
The epitopes for two monoclonal antibodies (MAbs) directed towards human apolipoprotein A-I (apoA-I), designated AI-1 and AI-3, have been more precisely defined. Previous work in our laboratory demonstrated that AI-1 and AI-3 recognize antigenic determinants located within cyanogen bromide (CNBr) fragments 1 (CF1) and 3 (CF3), respectively. Using peptides generated from endoproteinase cleavage of CF1 and CF3, we now report that both MAbs are specific for two previously unreported epitopes along the apoA-I molecule. The ability of whole endoproteinase digest mixtures to bind the MAbs, as determined by means of a competitive enzyme-linked immunosorbent assay (ELISA), indicated regions of CF1 and CF3 that were likely to form the epitopes. Purified peptides derived from the digests were then used to localize the epitopes recognized by MAbs AI-1 and AI-3 to within residues 28-47 and 140-147 of apoA-I, respectively. We have previously reported that the epitopes for both MAbs are exposed on HDL2, HDL3, and free apoA-I. Thus, the precise mapping of the binding sites recognized by AI-1 and AI-3 has enabled the identification of regions along apoA-I that are exposed on the surface of lipoprotein particles.
针对人载脂蛋白A-I（apoA-I）的两种单克隆抗体（MAb），即AI-1和AI-3的表位已得到更精确的定义。我们实验室之前的工作表明，AI-1和AI-3分别识别位于溴化氰（CNBr）片段1（CF1）和3（CF3）内的抗原决定簇。利用从CF1和CF3的内蛋白酶裂解产生的肽段，我们现在报告这两种单克隆抗体对apoA-I分子上两个以前未报道的表位具有特异性。通过竞争性酶联免疫吸附测定（ELISA）确定的完整内蛋白酶消化混合物与单克隆抗体结合的能力，表明了CF1和CF3中可能形成表位的区域。然后，将从消化物中纯化的肽段用于将单克隆抗体AI-1和AI-3识别的表位分别定位到apoA-I的28-47位残基和140-147位残基内。我们之前曾报道，这两种单克隆抗体的表位在HDL2、HDL3和游离apoA-I上均有暴露。因此，对AI-1和AI-3识别的结合位点的精确定位，使得能够确定apoA-I上暴露在脂蛋白颗粒表面的区域。
