Analysis of the function of human brain carboxypeptidase B and its diagnostic significance in dementia of Alzheimer's type

1.人脑羧肽酶B（HBCP B）的CPB-BP结合口袋的生物化学和计算机模拟分析：为了试图了解HBCP B及其特异性结合蛋白CPB-BP（1和2）的作用机制，我们通过使用基于两种蛋白质的一级结构产生的抗体开始了表位作图分析。这些单克隆抗体（6个克隆）表现出足够的特异性和灵敏度，并且对与CPB-BP（1和2）的结合及其调节的分析使得能够根据除了特异性和灵敏度之外的三维结构的差异对这些进行分类。这一成果为下一步实验方案中结合口袋和SBDD的结构分析奠定了基础.人血清中CPB-BP及其相关蛋白的研究为了促进用于此目的的HTP分析，利用表面等离子体相互作用分析与MALDI-TOF-MS的组合。使用对应于CPB的C-末端表位的肽配体，分析人外周血清样品，从其中去除主要的丰富蛋白质。除了先前通过免疫沉淀法确定的成功识别分子外，还识别了一种新分子，利用该分子识别与候选分子的相互作用。病理标本免疫组化分析。所有AD脑标本和部分正常老年脑标本海马均显示CPB-BP 2免疫反应阳性。

1. Biochemical and in silico analyses of the CPB-BP binding pocket of human brain carboxypeptidase B (HBCPB) : In an attempt to understand the mechanism of action of HBCPB and its specific binding protein, CPB-BP (1 & 2), we have started an epitope mapping analyses by using antibodies generated based upon primary structures of the both proteins. These monoclonal antibodies (6 clones) exhibit sufficient specificity and sensitivity, and analyses of binding to CPB-BP (1 & 2) and its regulation have enabled classification of these in terms of difference in three-dimensional structures, apart from specificity and sensitivity. This achievement embodies a basis for structural analysis of the binding pocket and SBDD in the next experimental program.2. A search and analysis for CPB-BP and related proteins in human serum. To facilitate HTP analyses for this purpose, the surface plasmon interaction analysis in combination with MALDI- TOF-MS was utilized. Using a peptide ligand corresponding to the C-terminal epitope of CPB, human peripheral serum samples, from which major abundant proteins were removed, were analyzed. In addition to the successful identification molecules previously determined by immunoprecipitation, a new molecule was identified, with which an interaction to the candidate molecule is recognized.3. Immunohistochemical analyses of pathological specimen. All AD brain samples and a portion of brain samples from the normal aged both exhibit CPB-BP2 immunoreactivity in the hippocampus.