A FRAMESHIFT MUTATION IN EXON V OF THE A-ALPHA-CHAIN GENE LEADING TO TRUNCATED A-ALPHA-CHAINS IN THE HOMOZYGOUS DYSFIBRINOGEN MILANO-III

An inherited dysfunctional fibrinogen variant, de noted as fibrinogen Milano III, was found in a 13-year-old girl suffering from recurrent v enous thrombosis, Plasma of the patient exhibited prolonged thrombin t ime and Reptilase time, Polymerization of fibrin monomers in the prese nce and absence of calcium ions was strongly impaired, SDS-polyacrylam ide gel electrophoresis of reduced fibrinogen showed normal B beta- an d gamma-chains, whereas no normal A alpha-chain was detected in the pr oposita. Immunoblot analysis with the monoclonal antibody Y18, detecti ng an epitope within the stretch of amino acids A alpha 1-51, revealed an A alpha-chain of about 50 kDa with an intact amino terminus. Immun oblotting with antibodies directed against serum albumin demonstrated the presence of albumin covalently linked to fibrinogen via a disulfid e bridge, The structural defect of fibrinogen Milano III was determine d by sequence analysis of a single-stranded fragment of genomic DNA am plified by polymerase chain reaction. An insertion of a thymine in the exon V of the A alpha-chain gene after the triplet ATT coding for Ile (A alpha 451) altered the reading frame and caused premature terminati on of the protein synthesis (Trp(452)(TGG)-Ser(453)(TCC)-Stop(454)(TGA )). In both parents, normal and mutant alleles were established, leadi ng to duplication of the sequence pattern after the thymine insertion site, whereas the proposita is homozygous for the new mutation in the fibrinogen A alpha-chain gene.