STRUCTURAL CHARACTERIZATION OF OLIGOSACCHARIDES AND UNDERSTANDING HEPARAN SULFATE-PROTEIN INTERACTIONS
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Hsieh, Po Hung. Structural Characterization Of Oligosaccharides And Understanding Heparan Sulfate-protein Interactions. 2016. https://doi.org/10.17615/yyyy-p788APA
Hsieh, P. (2016). STRUCTURAL CHARACTERIZATION OF OLIGOSACCHARIDES AND UNDERSTANDING HEPARAN SULFATE-PROTEIN INTERACTIONS. https://doi.org/10.17615/yyyy-p788Chicago
Hsieh, Po Hung. 2016. Structural Characterization Of Oligosaccharides And Understanding Heparan Sulfate-Protein Interactions. https://doi.org/10.17615/yyyy-p788- Last Modified
- March 19, 2019
- Creator
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Hsieh, Po-Hung
- Affiliation: Eshelman School of Pharmacy, Division of Chemical Biology and Medicinal Chemistry
- Abstract
- Heparin is known as a widely used anticoagulant drug since the 1930s. Heparin and heparan sulfate (HS) are highly sulfated polysaccharides that consist of a repeating disaccharide unit of glucosamine and glucuronic (GlcA) or iduronic acid (IdoA). The polydispersity of the saccharide sequence as well as the polyheterogeneity of the sulfation pattern make the study of this type of carbohydrate a formidable task. Instead of traditional extraction from animal mucus, we use the chemoenzymatic approach to synthesize homogeneous heparin oligosaccharides. The synthetic low molecular weight heparin was proven anti-FXa activity with reversible anticoagulant activity. In addition to secure the supply chain, the extra benefits aid future clinical usage of synthetic heparin drug. On the other side, the availability of synthetic pure common and less abundant HS oligosaccharides can serve as heparin component standards to secure the drug safety. Take the advantage of chemoenzymatic synthesis, the sufficient amount of structurally tailored oligosaccharide reagents are accessible. To investigate the interaction between HS and proteins, several oligosaccharide probes were successfully developed. Firstly, a synthesized heptasaccharide was used to co-crystalize with 2-O-sulfotransferase (2-OST) and probe the molecular basis of specificity. 2-OST was found to recognize N-sulfo and exclude 6-O-sulfo group of substrates, supporting the biosynthetic hypothesis 2-O-sulfation occurs after N-sulfation and prior to 6-O-sulfation. Secondly, we designed and synthesized the active heparan sulfate oligosaccharide probe carrying a diazoacetyl group. The resultant oligosaccharides demonstrate inhibitory activity and structural selectivity toward HS biosynthetic enzyme 2-OST and 3-OST, respectively. Thirdly, 13C-labeled saccharide was introduced at the desired site of the oligosaccharide to enhance the intensity of NMR signals. The labeled reagents facilitate directly analysis the binding between specific saccharide and protein. Finally, nuclear magnetic resonance analyses were performed to obtain coupling constants and full chemical shift assignments of all synthetic oligosaccharides. Comparing to relatively rigid GlcA, IdoA shows structural plasticity and diverse conformational preference in response to different sulfation patterns, which may be critical for various protein specific binding. The success of these projects assists the fundamental understand of HS-protein interaction as well as the design of next generation heparin-based therapeutics.
- Date of publication
- May 2016
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- Rights statement
- In Copyright
- Advisor
- Bowers, Albert
- Lee, Kuo-Hsiung
- Liu, Jian
- Jarstfer, Michael
- Pedersen, Lars
- Degree
- Doctor of Philosophy
- Degree granting institution
- University of North Carolina at Chapel Hill Graduate School
- Graduation year
- 2016
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