Marshall L B, Figler N L, Gonias S L
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville 22908.
J Biol Chem. 1992 Mar 25;267(9):6347-52.
Human alpha 2-macroglobulin (alpha 2M) exists in two well defined, highly distinct conformations and in less well described intermediate conformations. In this study, previously characterized reactions were used to partially or completely transform the conformation of alpha 2M. Electron micrographs of each preparation were subjected to image analysis. Ternary alpha 2M-trypsin (2 mol of trypsin/mol of alpha 2M) was analyzed as a control for the fully transformed state. Correspondence analysis (CORAN) and hierarchical ascendant classification (HAC) generated five image clusters from 330 aligned alpha 2M-trypsin complexes. Average images of each cluster resembled the letter "H" with four nearly equivalent lateral arms. Abnormally shaped lateral arms were not demonstrated by HAC, using a variety of factor sets. In a native polyacrylamide gel electrophoresis system, alpha 2M-thrombin migrated in a diffuse band partially behind alpha 2M-trypsin, suggesting conformational heterogeneity. CORAN and HAC of 733 alpha 2M-thrombin complexes identified two neighboring clusters, the average images of which showed an H-like structure in which one arm was replaced by a globular stain-excluding body. The two alpha 2M-thrombin clusters included 125 images (17.1% of image population). The complete absence of atypical lateral arm structure in the alpha 2M-trypsin clusters suggests that this variation is not the result of orientation or staining artifact. Native alpha 2M was reacted with cis-dichlorodiammineplatinum(II) and then with trypsin to form alpha 2M-Pt-trypsin, a preparation that includes partially transformed alpha 2M structures. CORAN and HAC of 580 alpha 2M-Pt-trypsin complexes generated five clusters, the average images of which showed atypical lateral arm structure equivalent to that demonstrated with alpha 2M-thrombin. The five alpha 2M-Pt-trypsin clusters accounted for 15.2% of the image population. These studies suggest that alpha 2M conformational change intermediates demonstrate common structural characteristics, permitting an elucidation of the steps involved in this complex transformation.
人α2-巨球蛋白（α2M）以两种明确且高度不同的构象以及一些描述较少的中间构象存在。在本研究中，利用先前已表征的反应来部分或完全转变α2M的构象。对每种制剂的电子显微照片进行图像分析。分析了三元α2M-胰蛋白酶（2摩尔胰蛋白酶/摩尔α2M）作为完全转变状态的对照。对应分析（CORAN）和层次上升分类（HAC）从330个对齐的α2M-胰蛋白酶复合物中生成了五个图像簇。每个簇的平均图像类似于字母“H”，有四个几乎相等的侧臂。使用各种因子集，HAC未显示出异常形状的侧臂。在天然聚丙烯酰胺凝胶电泳系统中，α2M-凝血酶在弥散条带中迁移，部分位于α2M-胰蛋白酶之后，表明构象异质性。对733个α2M-凝血酶复合物进行CORAN和HAC鉴定出两个相邻的簇，其平均图像显示出类似“H”的结构，其中一个臂被一个不染色的球状物体取代。这两个α2M-凝血酶簇包含125个图像（占图像总数的17.1%）。α2M-胰蛋白酶簇中完全没有非典型侧臂结构，这表明这种变化不是取向或染色假象的结果。天然α2M与顺式二氯二氨合铂（II）反应，然后与胰蛋白酶反应形成α2M-Pt-胰蛋白酶，该制剂包含部分转变的α2M结构。对580个α2M-Pt-胰蛋白酶复合物进行CORAN和HAC生成了五个簇，其平均图像显示出与α2M-凝血酶所显示的等效的非典型侧臂结构。这五个α2M-Pt-胰蛋白酶簇占图像总数的15.2%。这些研究表明，α2M构象变化中间体表现出共同的结构特征，从而能够阐明这一复杂转变所涉及的步骤。
