title: [The functional study of antithrombin L99 mutation].
abstract: OBJECTIVE
  To study the molecular mechanisms of inherited antithrombin (AT) deficiency caused by AT L99 mutation.
  
  
  METHODS
  Wild type (WT), L99V, L99A, L99I and L99S AT were purified from drosophila expression system. The binding capacity of AT and the low molecular weight heparin sodium was analyzed by the heparin binding assay. Surface plasmon resonance (SPR) was used to detect the binding ability of AT to thrombin (FIIa) or AT to coagulation factor Xa (FXa). The activity of AT(AT∶A)was detected by chromogenic assay.
  
  
  RESULTS
  The purified WT and mutant AT were at the same size. No additional band was observed by coomassie blue staining and western blot assay. Compared to the WT AT, the binding abilities of the low molecular weight heparin sodium to the AT L99V, L99A, L99I and L99S were (44.8±3.6)%, (118.9±14.0)%, (15.2±8.8)%, and(23.0±8.2)%, respectively. The binding abilities of FIIa to AT L99V, L99A, L99I and L99S were 13%, 57%, 3%, and 29%, while the binding of FXa to AT L99V, L99A, L99I and L99S were 7%, 51%, 1%, and 25%. The AT∶A of WT, L99V, L99A, L99I and L99S AT were 146.5%, 21.4%, 120.9%, 10.8%, and 39.0%, respectively.
  
  
  CONCLUSIONS
  The binding abilities of AT to heparin, FIIa and FXa were damaged by the L99 mutation, which resulted in decreased AT∶A and inherited AT deficiency.
authors.affiliation: Clinical Laboratory Department, Ruijin Hospital of Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
authors.forename: Tingting
authors.initials: T
authors.lastname: Yu
authors.affiliation: Clinical Laboratory Department, Ruijin Hospital of Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Dai
authors.affiliation: Clinical Laboratory Department, Ruijin Hospital of Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
authors.forename: Qiulan
authors.initials: Q
authors.lastname: Ding
authors.affiliation: Clinical Laboratory Department, Ruijin Hospital of Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
authors.forename: Qihua
authors.initials: Q
authors.lastname: Fu
authors.affiliation: Clinical Laboratory Department, Ruijin Hospital of Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Wang
mesh_terms: D000596:Amino Acids / Q000235:genetics*; D000818:Animals; D000990:Antithrombin III / Q000235:genetics; D020152:Antithrombin III Deficiency / Q000235:genetics*; D000991:Antithrombins; D004330:Drosophila; D015951:Factor Xa / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D009154:Mutation
chemical_list: D000596:Amino Acids; D000991:Antithrombins; D000990:Antithrombin III; D015951:Factor Xa