Molecular weight analysis of antithrombin III-heparin and antithrombin III-thrombin-heparin complexes

C. H. Pletcher, M. T. Cummingham, G. L. Nelsestuen

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The molecular interactions between components of the heparin-catalyzed antithrombin III/thrombin reaction were investigated by light scattering. When heparin was added to antithrombin III, the molecular weight increased to a maximum and then decreased to that of a 1:1 (antithrombin III·heparin) complex. The initial molecular weights at low heparin to antithrombin III ratios were consistent with the formation of a 2:1 (antithrombin III·heparin) complex in which only one antithrombin III molecule had undergone the conformational change measured by protein fluorescence enhancement. The peak molecular weight never reached that of complete 2:1 complex. This behavior was observed for bovine and human antithrombin III in the presence of both unfractionated heparin and high molecular weight-high affinity heparin. Pentosane polysulfate also caused some multiple associations. Bovine antithrombin III and thrombin formed a 1:1 complex that underwent further aggregation within minutes, while the human proteins did not aggregate on this time scale after forming the 1:1 complex. In the presence of stoichiometric amounts of heparin, the bovine proteins formed an initial complex of M(r) = 230,000 (corresponding to a dimer of heparin-antithrombin III-thrombin) which underwent further aggregation. The human proteins, however, formed a 1:1 (antithrombin III·thrombin) initial complex in the presence of heparin, followed by aggregation. These interactions of thrombin and antithrombin with heparin suggest complex interactions that could relate to heparin function.

Original languageEnglish (US)
Pages (from-to)4143-4147
Number of pages5
JournalJournal of Biological Chemistry
Issue number9
StatePublished - 1986


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