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Center for Nonlinear Studies |
Well-known for its role in regulating the coagulation cascade, heparin is a highly sulfated, flexible polysaccharide and one of the most negatively charged molecules in the human body, with an average net charge of -2.7 per disaccharide. Generated in mast cells, heparin has a large number of functions in the body that are mediated through interactions with proteins. As an anticoagulant, heparin interacts with antithrombin III (AT) in the coagulation cascade to prevent blood clotting. A single sequence of heparin (known as Arixtra) specifically activates AT causing a conformational change in its target protein, while other heparin sequences may also bind with much less activity. NMR studies have also been conducted on Arixtra, focusing on Arixtra’s chemical shifts and J3-coupling constants. However, a recent NMR study focused on determining Arixtra’s solution conformation by inferring intramolecular hydrogen bonding. With EXSY buildup curves and temperatures coefficients, Beecher et al. inferred the existence of specific intramolecular hydrogen bonds in Arixtra’s structure. The same study also conducted short MD simulations to rationalize these hydrogen bonds and found agreement between simulations and experiments. The same laboratory also investigated similar enoxaparin (low molecular weight heparin) sequences to Arixtra, investigating the effect of removing the 3O-sulfate on heparin’s conformation, since they showed that the 3O-sulfate forms key intramolecular hydrogen bonds in Arixtra. With insight on Arixtra’s experimental structure, it is highly useful to place this structure in the context of its conformational ensemble. The conformational ensemble provides thermodynamic and mechanistic details of heparin's stability, which may then be related to its biological activity. Accordingly, we are interested in using replica exchange molecular dynamics simulations in order to generate conformational ensembles of Arixtra in multiple force fields. After comparing these simulations to published NMR data, we can then conduct research on Arixtra’s sequence-conformation relationship. In order to do this, we intend to remove specific sulfates that are necessary for activity and then we intend to analyze differences within heparin’s conformational ensemble. As a result, these studies will allow us to establish the best available method for modeling heparin’s dynamics. Also, understanding heparin’s sequence-conformation relationship will provide insight on how to design heparin for a particular function, thus paving the way for a glycoengineering approach to heparin synthesis.
Host: Chris Neale