Project 3: Molecular interactions of marine carbohydrates and coagulation factors Junior Investigator: Vitor Pomin Mentors: Ikhlas Khan, University of Mississippi & Barbara Mulloy, Imperial College Cardiovascular diseases are the major cause of deaths not only in the USA, but also worldwide. Heparin is the most commonly used therapeutic. However, this sulfated glycan (SG) presents serious drawbacks such as bleeding trends, thrombocytopenia and contamination risks. Therefore, novel antithrombotic SGs with unique structures and different mechanisms of actions have been recently discovered, especially those from marine organisms. For example, a sulfated galactan reported by our group, isolated from the red alga Botryocladia occidentalis, shows (a) nearly equal in vivo action of unfractionated heparin (UFH) in arterial thrombosis, (b) equal in vivo effect of UFH in venous thrombosis at low doses (below 0.25 mg/kg of rat weight), (c) no bleeding risk at 5 and 10 mg/kg of rat weight, as opposed to the high-hemorrhagic behavior of UFH at the same doses, and (d) an in vitro anticoagulant mechanism of action distinct from UFH, with greater activity mediated through heparin cofactor II (HCII) while still retaining some activity through antithrombin. The IC50 value of the HCII-mediated anti-IIa activity of B. occidentalis sulfated galactan was around twenty times more active than the one from natural catalyst of HCII, dermatan sulfate. In order to fully understand the underlying mechanisms of action of this new marine sugar in the coagulation system, we will investigate at the atomic level the resultant intermolecular complexes made by HCII and thrombin with an oligosaccharide of defined structure isolated from the B. occidentalis sulfated galactan. We will also isolate, characterize, and screen the in vitro anticoagulant and in vivo antithrombotic activities of new SGs extracted from echinoderms collected in the Gulf of Mexico which were not previously investigated. Our laboratory is experienced solving the structures of the marine SGs using NMR along with chemical reactions and assessing their effects as potential anticoagulants and antithrombotics. Our latest results have demonstrated key structural requirements, such as 2-sulfation at galactose units, 4-sulfation at fucose units and longer chains for the anticoagulant and antithrombotic activities of the sea urchin-derived SGs. The marine sugars can serve as useful molecular tools in investigations of the different mechanism of action of sulfated glycans in coagulation and thrombosis. In- depth comprehension about these novel mechanisms can also facilitate the potential exploration of the marine sugars in the cardiovascular therapy. This research not only will identify potential medicinal molecules and elucidate their properties, but the combination of its results will also help to push the current status-quo of marine glycobiology towards a new level.
