A point mutation in a type I procollagen gene converts glycine 748 of the alpha 1 chain to cysteine and destabilizes the triple helix in a lethal variant of osteogenesis imperfecta.
Synthesis of type I procollagen was examined in fibroblasts from a proband with a lethal perinatal variant of osteogenesis imperfecta. After trypsin digestion of the type I procollagen, a portion of the alpha 1 (I) chains was recovered as disulfide-linked dimers. Digestion of the protein with vertebrate collagenase and mapping of cyanogen bromide peptides suggested that a new cysteine residue was present between residues 551 and 775 of the alpha 1 (I) chain. Sequencing of cloned cDNAs prepared using mRNA from the proband's fibroblasts demonstrated that some of the clones contained a single base mutation that converted the glycine codon in amino acid position 748 of the alpha 1 (I) chain to a cysteine codon. About 80% of the type I procollagen synthesized by the proband's fibroblasts had a decreased thermal stability. The results, therefore, were consistent with the conclusion that normal pro-alpha 1 (I) chains and pro-alpha 1 (I) chains containing a cysteine residue in the alpha chain domain were synthesized in about equal amounts and incorporated randomly into type I procollagen. However, only about 10% of the alpha 1 (I) chains generated by trypsin digestion were disulfide-linked. Further studies demonstrated a decreased rate of secretion of type I procollagen containing the new cysteine residue and decreased processing of the protein by procollagen N-proteinase in cultures of postconfluent fibroblasts. Both parents were phenotypically normal and their fibroblasts synthesized only normal type I procollagen. Therefore, the mutation in the proband was a sporadic one and is very likely to have caused the connective tissue fragility that produced the lethal phenotype.
Vogel
B E
BE
Department of Biochemistry and Molecular Biology, Jefferson Institute of Molecular Medicine, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Minor
R R
RR
Freund
M
M
Prockop
D J
DJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003545:Cysteine / Q000032:analysis*; D004247:DNA / Q000032:analysis; D004220:Disulfides; D005998:Glycine / Q000032:analysis*; D006358:Hot Temperature; D006801:Humans; D008297:Male; D009154:Mutation; D010013:Osteogenesis Imperfecta / Q000235:genetics* / Q000378:metabolism; D010455:Peptides / Q000032:analysis; D011347:Procollagen / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics*; D011487:Protein Conformation
D004220:Disulfides; D010455:Peptides; D011347:Procollagen; D004247:DNA; D003545:Cysteine; D005998:Glycine