A NOVEL AMYLOID COMPONENT

一种新型淀粉样蛋白成分

• 批准号: 6234021
• 负责人: TSUNAO SAITOH
• 金额: $ 15.65万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-04-01 至 1998-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6234021
• 关键词: Alzheimer's disease; amyloid proteins; amyloidosis; antisense nucleic acid; chromosomes; cytogenetics; gene expression; genetic promoter element; human genetic material tag; human tissue; immunocytochemistry; in situ hybridization; intermolecular interaction; laboratory mouse; laboratory rabbit; laboratory rat; messenger RNA; molecular pathology; northern blottings; nucleic acid sequence; protein isoforms; protein structure function; southern blotting; tissue /cell culture; western blottings

Amyloid accumulates as a core of senile plaques and along the vessel wall in the brain tissue of patients afflicted with Alzheimer's disease (AD). The major constituent of amyloid is a 39-43 amino acid peptide called amyloid Beta/A4-protein (A/Beta) derived from a precursor of around 100,000 daltons. Recent immunological studies have demonstrated many acute phase proteins are associated with amyloid, raising the possibility that amyloid might be composed of more than one type of protein. However, biochemically, Abeta had remained the only component identified for many years. However, recently, we have identified a new component in an AD amyloid biochemically and propose to study this protein using molecular biological, biochemical, immunological, and cell biological techniques. The complete digestion of purified AD amyloid, solubilized in 70% formic acid, and digested by cyanogen bromide and Achromobacter lyticus protease I yielded, in addition to ABeta fragments, two peptides of unknown sequence. These sequences were used to raise antisera. Two antisera portion of these sequences was used to generate oligonucleotide that were used by PCR to amplify DNA fragments from a brain cDNA library. A 247- nucleotide DNA segment was amplified. The sequencing of this short DNA demonstrated that this DNA encodes a protein that contains a perfect match with the peptide sequence found n amyloid. A full-length cDNA for the protein containing both two new amyloid sequence was isolated using this PCR amplified probe. We call the protein containing these peptide NACP for further immunohistochemical and immunochemical analysis of this protein. Antisense strategy will be used to identify the biological function of NACP. Finally, potential interaction between NAC or NACP and APP or Abeta will be investigated. Characterization of NAC and NACP should shed further light on the amyloidogenesis in AD and might help in finding the etiology of this devastating disease.

淀粉样蛋白作为老年斑的核心并沿着血管壁积聚 阿尔茨海默病（AD）患者的脑组织中。 淀粉样蛋白的主要成分是一种由 39-43 个氨基酸组成的肽，称为 淀粉样蛋白 Beta/A4 蛋白 (A/Beta) 源自周围的前体 100,000 道尔顿。 最近的免疫学研究表明许多 急性期蛋白与淀粉样蛋白相关，增加了可能性 淀粉样蛋白可能由不止一种类型的蛋白质组成。 然而，从生化角度来看，Abeta 仍然是唯一被识别的成分 多年。 然而，最近我们发现了一个新组件 AD 淀粉样蛋白的生化研究，并建议使用 分子生物学、生物化学、免疫学和细胞生物学 技术。 完全消化纯化的 AD 淀粉样蛋白，溶解在 70% 甲酸中 酸，并被溴化氰和无色杆菌蛋白酶消化 除了 Aβ 片段之外，我还得到了两种未知的肽 顺序。 这些序列用于产生抗血清。 两份抗血清 这些序列的一部分用于生成寡核苷酸 PCR 用于扩增大脑 cDNA 文库中的 DNA 片段。 247- 核苷酸DNA片段被扩增。 这段短DNA的测序 证明该 DNA 编码的蛋白质含有完美的 与淀粉样蛋白中发现的肽序列相匹配。 全长 cDNA 使用以下方法分离含有两个新淀粉样蛋白序列的蛋白质 该PCR扩增探针。 我们将含有这些肽的蛋白质称为 NACP 用于进一步免疫组织化学和免疫化学分析 蛋白质。 反义策略将用于鉴定生物 NACP 的功能。 最后，NAC 或 NACP 与 APP或Abeta将被调查。 NAC 和 NACP 的表征 应该进一步阐明 AD 中的淀粉样蛋白生成，并可能有助于 找到这种毁灭性疾病的病因。