Marriott A, Ayad S, Grant M E
Department of Biochemistry and Molecular Biology, School of Biological Sciences, University of Manchester, UK.
J Cell Sci. 1991 Jul;99 ( Pt 3):641-9. doi: 10.1242/jcs.99.3.641.
Chondrocytes were isolated from bovine growth-plate cartilage and cultured within type I collagen gels. A major collagen with chains of Mr 59,000, decreasing to 47,000 on pepsinization, was synthesized and identified as type X collagen. This collagen was cleaved at two sites by mammalian collagenase, resulting in a major triple-helical fragment with chains of Mr 32,000. The species of Mr 59,000, 47,000 and 32,000 were not detected by SDS-polyacrylamide gel electrophoresis before reduction, indicating the presence of disulphide bonds within the triple helix. In contrast, similar biosynthetic studies with human growth-plate cartilage in organ culture, indicated that human type X collagen does not contain disulphide bonds. A polyclonal antiserum was raised to bovine type X collagen and used in immunolocalization studies to provide direct evidence for the association of type X collagen with the hypertrophic chondrocytes in both bovine and human growth plates during development.
从牛生长板软骨中分离出软骨细胞，并在I型胶原凝胶中培养。合成了一种主要的胶原蛋白，其链的分子量为59,000，经胃蛋白酶处理后降至47,000，并被鉴定为X型胶原。这种胶原蛋白被哺乳动物胶原酶在两个位点切割，产生一个主要的三螺旋片段，其链的分子量为32,000。在还原前，通过SDS-聚丙烯酰胺凝胶电泳未检测到分子量为59,000、47,000和32,000的条带，这表明三螺旋内存在二硫键。相比之下，对器官培养中的人生长板软骨进行的类似生物合成研究表明，人X型胶原不含有二硫键。制备了针对牛X型胶原的多克隆抗血清，并用于免疫定位研究，以提供直接证据证明在发育过程中，X型胶原与牛和人生长板中的肥大软骨细胞相关联。
