Research Institute for Biological Sciences, Tokyo University of Science, 2669, Yamazaki, Noda, Chiba 278-0022, Japan.
J Biochem. 2011 May;149(5):569-80. doi: 10.1093/jb/mvr007. Epub 2011 Jan 21.
Monoclonal antibodies (mAbs) were prepared to analyse the conformation of human serum albumin (HSA) and its non-enzymatic glycation (NEG) products. We first determined the epitopes of the mAbs using HSA subdomains expressed on the surface of yeast. Each mAb was classified as belonging to one of two groups; Type I mAbs which recognized a single subdomain structure and Type II mAbs which bound to plural subdomains. We analysed the pH-dependent conformational change in HSA. We found that one Type II mAb, HAy2, detected the normal to base form (N-B) transition while the other did not, suggesting that N-B transition occurred around Domain I accompanied by topological isomerization of subdomains without changing the subdomain structure itself. Next, we analysed the conformations of the NEG products. Since all mAbs reacted with the early NEG products, no structural change was thought to have occurred in the early NEG products. On the other hand, only a Type I mAb, HAy1, had full binding activity with the advanced glycation end products (AGE) while the other mAbs had lost or had diminished activity, suggesting that the over-all tertiary structure of HSA was altered except for a subdomain, sDOM Ia, in AGE.
单克隆抗体（mAbs）被制备用于分析人血清白蛋白（HSA）及其非酶糖基化（NEG）产物的构象。我们首先使用在酵母表面表达的 HSA 亚域来确定 mAb 的表位。每种 mAb 都被归类为属于两种类型之一；识别单个亚域结构的 I 型 mAb 和结合多个亚域的 II 型 mAb。我们分析了 HSA 中 pH 依赖性的构象变化。我们发现，一种 II 型 mAb，HAy2，检测到正常到碱形式（N-B）转变，而另一种则没有，这表明 N-B 转变发生在 I 结构域周围，伴随着亚域的拓扑异构化，而不改变亚域结构本身。接下来，我们分析了 NEG 产物的构象。由于所有 mAb 都与早期 NEG 产物反应，因此认为早期 NEG 产物没有发生结构变化。另一方面，只有一种 I 型 mAb，HAy1，与晚期糖基化终产物（AGE）具有完全结合活性，而其他 mAb 则失去或活性降低，这表明除了 AGE 中的 sDOM Ia 亚域外，HSA 的整体三级结构发生了改变。
