A substitution of cysteine for glycine 748 of the alpha 1 chain produces a kink at this site in the procollagen I molecule and an altered N-proteinase cleavage site over 225 nm away.
A substitution of cysteine for glycine 748 of the alpha 1 chain produces a kink at this site in the procollagen I molecule and an altered N-proteinase cleavage site over 225 nm away.
复制标题
DOI：
10.1016/s0021-9258(18)37416-7
复制
发表时间：
1988-12
期刊：
影响因子：
--
通讯作者：
B. Vogel;R. Doelz;K. Kadler;Y. Hojima;J. Engel;D. Prockop
中科院分区：
文献类型：
--
作者：
B. Vogel;R. Doelz;K. Kadler;Y. Hojima;J. Engel;D. Prockop
在以前的工作（Vogel，B. E、迈纳河R.，Freund，M.，和Prockop，D. J.（1987）J.Biol.Chem.262，14737-14744），我们鉴定了一个单碱基突变，该突变在具有骨生成突变的致死变体的先证者中将I型前胶原的α 1链的第748位的甘氨酸转化为半胱氨酸。除了翻译后过度修饰，异常分子显示出降低的热稳定性和降低的分泌速率。一个无法解释的发现是，前胶原在皮肤成纤维细胞融合后培养物中难以加工成pCcollagen。在这里，我们表明，由先证者的细胞合成的前胶原是抵抗裂解的前胶原N-蛋白酶，构象敏感的酶。由于分子中唯一可检测到的缺陷是甘氨酸取代的半胱氨酸，我们组装了几个空间填充模型，试图解释超过700个氨基酸残基或225 nm的氨基酸取代如何影响N-蛋白酶切割位点的结构。该模型在一条或两条α 1链中掺入了三肽单元的相移。最令人满意的模型在半胱氨酸取代位点产生30度或60度的柔性扭结。因此，我们用电子显微镜检查了前胶原。约25%的分子具有在对照样品中未观察到的扭结，并且扭结位于半胱氨酸取代的位点。
In previous work (Vogel, B. E., Minor, R. R., Freund, M., and Prockop, D. J. (1987) J. Biol. Chem. 262, 14737-14744), we identified a single-base mutation that converted the glycine at position 748 of the alpha 1 chain of type I procollagen to a cysteine in a proband with a lethal variant of osteogenesis imperfecta. In addition to posttranslational overmodification, the abnormal molecules displayed decreased thermal stability and a decreased rate of secretion. An unexplained finding was that procollagen was poorly processed to pCcollagen in postconfluent cultures of skin fibroblasts. Here, we show that the procollagen synthesized by the proband's cells is resistant to cleavage by procollagen N-proteinase, a conformation-sensitive enzyme. Since the only detectable defect in the molecule was the cysteine for glycine substitution, we assembled several space-filling models to try to explain how the structure of the N-proteinase cleavage site can be affected by an amino acid substitution over 700 amino acid residues or 225 nm away. The models incorporated a phase shift of a tripeptide unit in one or both of the alpha 1 chains. The most satisfactory models produced a flexible kink of 30 degrees or 60 degrees at the site of the cysteine substitution. Therefore, we examined the procollagen by electron microscopy. About 25% of the molecules had a kink not seen in control samples, and the kink was at the site of the cysteine substitution.