SUBSTITUTION OF ASPARTIC-ACID FOR GLYCINE AT POSITION-310 IN TYPE-II COLLAGEN PRODUCES ACHONDROGENESIS-II, AND SUBSTITUTION OF SERINE AT POSITION-805 PRODUCES HYPOCHONDROGENESIS - ANALYSIS OF GENOTYPE-PHENOTYPE RELATIONSHIPS

Two different mutations were found in two unrelated probands with leth al chondrodysplasias, one with achondrogenesis type II and the other w ith the less severe phenotype of hypo-chondrogenesis. The mutations in the COL2A1 gene were identified by denaturing gradient gel electropho resis analysis of genomic DNA followed by dideoxynucleotide sequencing and restriction site analysis. The proband with achondrogenesis type II had a heterozygous single-base mutation that substituted aspartate for glycine at position 310 of the alpha 1(II) chain of type II procol lagen, The proband with hypochondrogenesis had a heterozygous single-b ase mutation that substituted serine for glycine at position 805. Type II collagen extracted from cartilage from the probands demonstrated t he presence of type I collagen and a delayed electrophoretic mobility, indicating post-translational overmodifications. Analysis of CNBr pep tides showed that, in proband 1, the entire peptides were overmodified . Examination of chondrocytes cultured in agarose or alginate indicate d that there was a delayed secretion of type II procollagen. In additi on, type II collagen synthesized by cartilage fragments from the proba nds demonstrated a decreased thermal stability. The melting temperatur e of the type II collagen containing the aspartate-for-glycine substit ution was reduced by 4 degrees C, and that of the collagen containing the serine-for-glycine substitution was reduced by 2 degrees C. Electr on microscopy of the extracellular matrix from the chondrocyte culture s showed a decreased density of matrix and the presence of unusually s hort and thin fibrils. Our results indicate that glycine substitutions in the N-terminal region of the type II collagen molecule can produce more severe phenotypes than mutations in the C-terminal region, The a spartate-for-glycine substitution at position 310, which was associate d with defective secretion and a probable increased degradation of col lagen, is the most destabilizing mutation yet reported in type II proc ollagen.