heparin  dodecasaccharide   3D structure of IL-8 dimer   If the plug-in named ChemScape Chime is properly installed on your computer, you should see a rotating 3D heparin molecule. The molecule can be moved by clicking it with the left button of your mouse and its characteristics, including automatic rotation, can be modified by clicking it with the right button.  To download the plug-in: .      Back to Presta's lab

ABSTRACT Tat protein, a transactivating factor of the human immunodeficiency virus type 1, acts also as an extracellular molecule, modulating gene expression, cell survival, growth, transformation, and angiogenesis. Heparin affects the bioavailability and biological activity of extracellular Tat (Rusnati, M., Coltrini, D., Oreste, P., Zoppetti, G., Albini, A., Noonan, D., D’Adda di Fagagna, F., Giacca, M., and Presta, M. (1997) J. Biol. Chem. 272, 11313-11320). Here, a series of homogeneously sized, [3H]-labeled heparin fragments were evaluated for their capacity to bind to free glutathione-S-transferase (GST)-Tat protein and to GST-Tat immobilized to glutathione-agarose resin beads.  The results demonstrate that hexasaccharides represent the minimal sized heparin fragments able to interact with GST-Tat at physiological ionic strength. Also, the affinity of binding increases with increasing the molecular size of the oligosaccharides, large fragments (> 18-saccharides) approaching the affinity of full-size heparin. Scatchard plot analysis of the binding data indicates that 6-mer heparin binds GST-Tat with a dissociation constant (Kd) equal to 0.7 microM and a molar oligosaccharide:GST-Tat ratio of about 1:1. Interaction of GST-Tat with 22-mer or full-size heparin is consistent instead with a two component binding: at sub-saturating concentrations, a single molecule of heparin interacts with 4-6 molecules of GST-Tat with high affinity (Kd values in the nM range of concentration); at saturating concentrations, heparin binds GST-Tat with lower affinity (Kd values in the microM range of concentration) and a molar oligosaccharide:GST-Tat ratio of about 1:1.  In agreement with the binding data, a positive correlation exists between the size of heparin oligosaccharides and their capacity to inhibit cell internalization and LTR-transactivating activity of extracellular Tat in HL3T1 cells and its mitogenic activity in murine adenocarcinoma T53 Tat-less cells. In all the assays, at least 18-20 saccharide residues are required to confer to the oligosaccharide chain an antagonist potency similar to that of full-size heparin. The data demonstrate that the modality of heparin-Tat interaction is strongly affected by the size of the saccharide chain and by the relative concentration of the two molecules. The possibility to establish multiple interactions increases the affinity of large heparin fragments for Tat protein and the capacity of the glycosaminoglycan to modulate the biological activity of extracellular Tat.       J. Biol. Chem. 1999 274 :28198-28205.

AIRC: Special Project Angiogenesis