ELECTRON MICROSCOPY OF HUMAN ALPHA-2-MACROGLOBULIN

人 ALPHA-2-巨球蛋白的电子显微镜

• 批准号: 3361239
• 负责人: JAMES K STOOPS
• 金额: $ 11.62万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-09-01 至 1993-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3361239
• 关键词: chemical binding; cryoscience; glycoprotein structure; human tissue; image enhancement; immunoelectron microscopy; immunoglobulins; macroglobulins; protease inhibitor; protein structure function; receptor binding; stainings; thioester; trypsin

alpha2-Macroglobulin (alpha2M) is a major inhibitor of virtually all known endoproteases found in the plasma of vertebrates. It has been suggested that it serves as a general protease scavenger, thereby protecting the blood and tissue proteins from degradation. As a protease inhibitor, it may play a role in the regulation of protease activity in fibrinolysis, coagulation, and complement activation. As a result, alpha2M is a large glycoprotein (Mr=725,000) composed of four identical subunits. Protease inhibition is accompanied by a major change in its structure resulting in entrapment of the protease. In order to determine the mechanism by which the native structure rearranges to the protease-bound form, immunoelectron microscopy will be employed. The structures will be visualized using both negative-stain and cryoelectron microscopy and maximum resolution of these micrographs will be achieved by image processing. Immunoelectron microscopy and gold cluster labeling will also be utilized to locate the sites of biological interest, which include the "bait" region, the receptor-binding sites, and the thioesters. Further details of the structural consequences of protease binding will be studied by electron microscopy of the methylamine-treated and the alpha2M-protease complexes with one and two molecules of trypsin bound.

α2-巨球蛋白(α2M)是几乎所有已知的 脊椎动物血浆中发现的内切酶。有人建议说， 它作为一种普通的蛋白酶清除剂，从而保护 血液和组织蛋白质的降解。作为一种蛋白酶抑制剂，它 可能在调节纤溶过程中的蛋白酶活性中发挥作用， 凝血和补体激活。因此，alpha2M是一个很大的 糖蛋白(Mr=725,000)由四个相同的亚基组成。纤维素酶 抑制伴随着其结构的重大变化，导致 蛋白水解酶的包埋。为了确定所依据的机制 天然结构重新排列为与蛋白酶结合的形式，免疫电子 将使用显微技术。这些结构将使用这两个工具可视化 负染和冷冻电子显微镜及其最大分辨率 显微摄影将通过图像处理来实现。免疫电子 显微镜和金簇标记也将被用来定位 具有生物价值的地点，其中包括“诱饵”区域、 受体结合部位和硫代酯。更详细的信息 将用电子学研究蛋白酶结合的结构后果 甲胺处理和α2M-蛋白酶复合体的显微镜观察 结合了一个和两个胰酶分子。