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      Requirement of lysine residues outside of the proposed pentasaccharide binding region for high affinity heparin binding and activation of human antithrombin III.

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      The Journal of biological chemistry

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          Abstract

          Variant forms of human antithrombin III with glutamine or threonine substitutions at Lys114, Lys125, Lys133, Lys136, and Lys139 were expressed in insect cells to evaluate their roles in heparin binding and activation. Recombinant native ATIII and all of the variants had very similar second order rate constants for thrombin inhibition in the absence of heparin, ranging from 1.13 x 10(5) M-1min-1 to 1.66 x 10(5) M-1min-1. Direct binding studies using 125I-flouresceinamine-heparin yielded a Kd of 6 nM for the recombinant native ATIII and K136T, whereas K114Q and K139Q bound heparin so poorly that a Kd could not be determined. K125Q had a moderately reduced affinity. Heparin binding affinity correlated directly with heparin cofactor activity. Recombinant native ATIII was nearly identical to plasma-purified ATIII, whereas K114Q and K139Q were severely impaired in heparin cofactor activity. K125Q and K136T were only slightly impaired. Based on these data, Lys114 and Lys139, which are outside of the putative pentasaccharide binding site, play pivotal roles in the high affinity binding of heparin to ATIII and the activation of thrombin inhibitory activity.

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          Author and article information

          Journal
          J. Biol. Chem.
          The Journal of biological chemistry
          0021-9258
          0021-9258
          Aug 23 1996
          : 271
          : 34
          Affiliations
          [1 ] Department of Developmental and Cell Biology, School of Biological Sciences, University of California, Irvine, California 92717, USA.
          Article
          8702852
          636e8ae7-5310-4117-813e-3d59e45cb5fe
          History

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