A Gly421->Asp substitution causes unusual electrophoretic mobility ofα2(I) chain in a lethal case of Osteogenesis Imperfecta. † 602

Abstract

Biochemical analysis of type I collagen synthesized by cultured skin fibroblasts was performed to characterized the defect in a patient affected by lethal Osteogenesis Imperfecta (OI), a dominant heritable brittle bone disorder caused by mutations in one of the two genes coding for the α1 and α2 chains of type I collagen. The SDS-urea-PAGE of procollagen and collagen revealed a broad α1(I) chain, a normal α2(I) and anα2(I) migrating half way between α1 and α2. Procollagen and collagen of media and cell layers, synthesized in the presence ofαα′-dipyridyl, revealed both normal and slower α2(I) suggesting an insertion in COL1A2 gene. CNBr peptide digestion of collagen yielded overmodified α1(I) CB3 and CB7 peptides and a delayed migration of the α2(I) CB3-5 peptide. A delayed CB3-5 persisted also afterαα′-dipyridyl treatment, supporting an alteration ofα2(I) size. The mutation was localized, by this data, between aa 353-551 in α2(I) (CB 3-5). The sequence of this region revealed a G->A transition at nt 1671 in one allele, changing Gly421->Asp in oneα2(I) chain. Procollagen processing was normal. The Tm of the slowα2(I) collagen was 2iC lower than the control, indicating a decrease in triple helix stability. The mutant protein was incorporated in the extracellular matrix synthesized by long term cultured fibroblasts.