Mutation analysis of ITGB3 showed that a novel mutation F153S (F179S) in exon 4 and an intronic mutation IVS5(+1) G>A are located on different alleles. (A) Electropherograms of gDNA sequencing reveal a previously reported splice site mutation in intron 5: IVS5(+1) G>A (left) and a novel missense mutation in exon 4 c.556T>C encoding F153S (F179S numbering with cleaved propeptide) (right). The precise nucleotide of each mutation is indicated by a heavy arrow (n = 2 in triplicate with forward and reverse primers). (B) Platelet cDNA sequencing analysis detected only 1 transcript with ITGB3 mutation: c.556 T>C (F153S or F179S). Sequence analysis primary cDNA (n = 1 in triplicate using forward and reverse primers). The cDNA PCR products were also cloned into a plasmid and sequence analysis confirmed GT hemizygous expression (n = 20 clones using forward and reverse primers). (C) A molecular ribbon diagram designed with PyMol software depicting an extended conformation of αIIbβ3 generated from the crystal structure in the PDB 3fcs shows the location of F153Sβ3 as green sticks and dots.31 Metal ions are shown as blue spheres. Disulfide bonds are shown as yellow sticks.
