Constantinou C D, Nielsen K B, Prockop D J
Department of Biochemistry and Molecular Biology, Jefferson Institute of Molecular Medicine, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
J Clin Invest. 1989 Feb;83(2):574-84. doi: 10.1172/JCI113920.
A fraction of the pro alpha 1(I) and pro alpha 2(I) chains in type I procollagen synthesized by the fibroblasts from a proband with a lethal variant of osteogenesis imperfecta were overmodified by posttranslational reactions. After digestion with pepsin, some of the alpha 1(I) chains were recovered as disulfide-linked dimers. Mapping of cyanogen bromide peptides indicated that the disulfide link was contained in alpha 1-CB6, the cyanogen bromide fragment containing amino acid residues 823-1014 of the alpha 1(I) chain. Nucleotide sequencing of cDNA clones demonstrated a substitution of T for G that converted glycine 904 of the alpha 1(I) chain to cysteine. A large fraction of the type I procollagen synthesized by the proband's fibroblasts had a thermostability that was 3-4 degrees C lower than the normal type I procollagen as assayed by brief proteinase digestion. In addition, the type I procollagen synthesized by the proband's fibroblasts was secreted with an abnormal kinetic pattern in that there was a lag period of about 30 min in pulse-chase experiments. The mutation of glycine to cysteine was not found in type I procollagen synthesized by fibroblasts from the proband's parents. Therefore, the mutation was a sporadic one. However, the mother's fibroblasts synthesized a type I procollagen in which part of the pro alpha chains were overmodified and had a lower thermostability. Therefore, the proband may have inherited a mutated allele for type I procollagen from her mother that contributed to the lethal phenotype. The mother was asymptomatic. She was somewhat short and had slightly blue sclerae but no definitive signs of a connective tissue abnormality. The observations on the mother indicated, therefore, that a mutation that causes synthesis of a type I procollagen with a lowered thermal stability does not necessarily produce a heritable disorder of connective tissue.
一名患有致死性成骨不全变异型的先证者的成纤维细胞合成的I型前胶原中,一部分前α1(I)链和前α2(I)链通过翻译后反应过度修饰。用胃蛋白酶消化后,一些α1(I)链以二硫键连接的二聚体形式回收。溴化氰肽图谱表明,二硫键存在于α1-CB6中,α1-CB6是溴化氰片段,包含α1(I)链的氨基酸残基823 - 1014。cDNA克隆的核苷酸测序显示,T替代了G,导致α1(I)链的甘氨酸904转变为半胱氨酸。通过短暂蛋白酶消化测定,先证者的成纤维细胞合成的大部分I型前胶原的热稳定性比正常I型前胶原低3 - 4℃。此外,先证者的成纤维细胞合成的I型前胶原以异常的动力学模式分泌,在脉冲追踪实验中有大约30分钟的延迟期。在先证者父母的成纤维细胞合成的I型前胶原中未发现甘氨酸突变为半胱氨酸的情况。因此,该突变是散发性的。然而,母亲的成纤维细胞合成的I型前胶原中,部分前α链过度修饰且热稳定性较低。因此,先证者可能从母亲那里遗传了一个I型前胶原的突变等位基因,这导致了致死表型。母亲无症状。她身材稍矮且巩膜略呈蓝色,但没有明确的结缔组织异常体征。因此,对母亲的观察表明,导致合成热稳定性降低的I型前胶原的突变不一定会产生遗传性结缔组织疾病。