Identification of a novel amino acid deletion mutation and a very rare single nucleotide variant in a Japanese family with type I antithrombin deficiency

We studied a Japanese family with type I antithrombin (AT) deficiency and identified a novel in-frame deletion mutation (-ATG at nucleotide position of 2771-2773) in the AT gene, which predicted loss of a methionine (Met) at amino acid number of 103. In addition, we found a single base replacement of G to A at nucleotide position of 67 (4 base upstream to the initial codon) in the mutant allele. Since the G67A substitution in the AT gene was very rare, this family was the second case, in which the nucleotide change was transmitted. To elucidate the mechanism of AT deficiency, we transiently expressed wild type and the mutant AT (ΔM103) in HuH-7 human hepatoma cells and performed pulse-chase studies. The experiments revealed that the mutant AT (ΔM103) hardly secreted into the medium and underwent partial intracellular degradation. In addition, we performed luciferase reporter assay to examine the effect of G67A substitution on the AT gene expression, and found that the substitution did not reduce the luciferase activity. These results suggested that secretion defect and intracellular degradation of the variant molecule with the deletion of Met 103 were responsible for AT deficiency in this family.