The presence of intermolecular disulfide cross-links in type III collagen.
The presence of intermolecular disulfide cross-links in type III collagen.
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DOI：
10.1016/s0021-9258(18)32246-4
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发表时间：
1983-06
期刊：
影响因子：
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通讯作者：
D. Cheung;P. Dicesare;P. Benya;E. Libaw;M. Nimni
中科院分区：
文献类型：
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作者：
D. Cheung;P. Dicesare;P. Benya;E. Libaw;M. Nimni
分析了牛和大鼠III型胶原制剂中分子间二硫交联的存在和体外形成。利用胃蛋白酶从胎牛皮肤中提取III型胶原蛋白，并通过差异盐沉淀法、胍变性法和变性法纯化。几乎所有的III型胶原都以还原敏感的γ -组分和高分子量的聚集体的形式存在。用CNBr切割后，用二维图谱分析肽段。羧基末端CNBr肽CB9B的六聚体证明了分子间二硫键的存在。该交联肽通过还原完全转化为CB9B单体。采用β -氨基本体腈处理大鼠皮肤的III型胶原蛋白，检测其分子间二硫交联的体外形成。这是由盐提取，差异盐沉淀和胃蛋白酶处理制备的。十二烷基硫酸钠凝胶电泳在部分纯化的III型胶原蛋白重组成纤维之前主要检测到γ链。在重组成纤维后，大部分材料以高分子量聚集体的形式存在。还原后，这些聚集体主要产生α链。这些数据证明了天然III型胶原蛋白分子间二硫键的存在及其在体外纤维形成过程中的形成。
Bovine and lathyritic rat type III collagen preparations were analyzed for the presence and in vitro formation of intermolecular disulfide cross-links. Type III collagen from fetal bovine skin was extracted with the aid of pepsin and purified by differential salt precipitation, guanidine denaturation, and renaturation. Nearly all of the type III collagen was present as reduction-sensitive gamma-components and higher molecular weight aggregates. After cleavage with CNBr, the peptides were analyzed by two-dimensional mapping. The presence of intermolecular disulfide bonds was demonstrated by the existence of a hexamer of the COOH-terminal CNBr peptide, CB9B. This cross-linked peptide was completely converted to the CB9B monomer by reduction. Type III collagen from the skins of beta-aminoproprionitrile-treated rats was used to test for the in vitro formation of intermolecular disulfide cross-links. This was prepared by salt extraction, differential salt precipitation, and pepsin treatment. Sodium dodecyl sulfate-gel electrophoresis of this partially purified type III collagen before reconstitution into fibers detected primarily gamma-chains. After reconstitution into fibers, the majority of the material was present as higher molecular weight aggregates. Upon reduction, these aggregates generated predominantly alpha-chains. These data demonstrate the existence of intermolecular disulfide bonds in native type III collagen and their formation during in vitro fibrillogenesis.