THROMBOEMBOLIC DISEASE DUE TO THERMOLABILE CONFORMATIONAL-CHANGES OF ANTITHROMBIN ROUEN-VI (187-ASN-]ASP)

被引:147
作者
BRUCE, D
PERRY, DJ
BORG, JY
CARRELL, RW
WARDELL, MR
机构
[1] UNIV CAMBRIDGE, MRC CTR, DEPT HAEMATOL, CAMBRIDGE CB2 2QH, ENGLAND
[2] HOP CHARLES NICOLLE, HEMATOL LAB, F-76031 ROUEN, FRANCE
基金
英国惠康基金;
关键词
LATENT-ANTITHROMBIN; SERPINS; LOOP-SHEET POLYMERIZATION; ANTITHROMBIN III; PROTEIN CONFORMATION;
D O I
10.1172/JCI117589
中图分类号
R-3 [医学研究方法]; R3 [基础医学];
学科分类号
1001 ;
摘要
A new variant of antithrombin (Rouen-VI, 187 Asn-->Asp) with increased heparin affinity was shown to have normal inhibitory activity which decreased slowly at 4 degrees C and rapidly at 41 degrees C. On electrophoresis the freshly isolated variant had an anodal shift relative to native antithrombin due to the mutation. A further anodal transition occurred after either prolonged storage at 4 degrees C or incubation at 41 degrees C due to the formation of a new inactive uncleaved component with properties characteristic of L-form (latent) antithrombin. At the same time, polymerization also occurred with a predominance of di-, tri-, and tetra-mers. These findings fit with the observed mutation of the conserved asparagine (187) in the F-helix destabilizing the underlying A-sheet of the molecule. Evidence of A-sheet perturbation is provided by the increased rate of peptide insertion into the A-sheet and by the decreased vulnerability of the reactive loop to proteolysis. The spontaneous formation of both L-antithrombin and polymers is consistent with our crystal structure of intact antithrombin where L-form and active antithrombin are linked together as dimers. The nature of this linkage favors a mechanism of polymerization whereby the opening of the A-sheet, to give incorporation of the reactive center loop, is accompanied by the bonding of the loop of one molecule to the C-sheet of the next. The accelerated lability of antithrombin Rouen-VI at 41 versus 37 degrees C provides an explanation for the clinical observation that episodes of thrombosis were preceded by unrelated pyrexias.
引用
收藏
页码:2265 / 2274
页数:10
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