title: [Molecular mechanisms of antithrombin deficiency caused by novel double heterozygous mutations].
abstract: OBJECTIVE
  To explore the molecular mechanisms of antithrombin (AT) deficiency caused by novel double heterozygous mutations.
  
  
  METHODS
  Wild-type and mutant AT cDNA expression plasmids (ATwt, AT-c.134G > A, AT-c.342T > G, AT-c.134G > A and AT-c.342T > G) were transfected into HEK293T cells. Western blot was used to detect the AT:Ag in cell lysates. Homology was used to reestablish 3-D spatial structure of AT. Laser confocal assay was utilized to analyze the distribution of AT in endoplasmic reticulum (ER).
  
  
  RESULTS
  Compared to the wild-types, the AT expression of HEK293T cells sharply increased when they were transfected by AT-c.342T > G or AT-c.134G > A and c.342T > G. Homology modeling showed that the mutation (AT-c.342T > G) caused a decreased distance between Arg and surrounding bases as Arg's side chain was significantly longer than Ser's. The mutation of 13th base pair decreases the distance between AT and heparin. Laser confocal assay showed that the AT protein concreted in HEK293T cells when they were transfected by mutant plasmids (AT-c.134G > A and c.342T > G) and aggregated in ER.
  
  
  CONCLUSIONS
  The main molecular mechanism of AT deficiency in this pedigree is the disturbed AT secretion as a result of the mutation of AT-c.342T > G.
authors.affiliation: Department of Vascular Surgery, Shanghai JiaoTong University, Shanghai, China.
authors.forename: Hao-yu
authors.initials: HY
authors.lastname: Deng
authors.forename: Jia-quan
authors.initials: JQ
authors.lastname: Chen
authors.forename: Hui
authors.initials: H
authors.lastname: Xie
authors.forename: Yi
authors.initials: Y
authors.lastname: Gu
authors.forename: Ji-wei
authors.initials: JW
authors.lastname: Zhang
authors.forename: Hao
authors.initials: H
authors.lastname: Zhang
authors.forename: Zhao-xiong
authors.initials: ZX
authors.lastname: Zhou
authors.forename: Wei
authors.initials: W
authors.lastname: Liang
authors.forename: Guan-hua
authors.initials: GH
authors.lastname: Xue
authors.forename: Peng
authors.initials: P
authors.lastname: Wang
authors.forename: Lan
authors.initials: L
authors.lastname: Zhang
mesh_terms: D020152:Antithrombin III Deficiency / Q000235:genetics*; D057809:HEK293 Cells; D006579:Heterozygote; D006801:Humans; D009154:Mutation; D010957:Plasmids; D014162:Transfection; D054556:Venous Thromboembolism / Q000235:genetics*