Heather Bethea, a graduate student in the Division of Medicinal Chemistry and Natural Products, will defend her dissertation on Thursday, March 25.

Bethea's dissertation is titled "Structural and Functional Analysis of Heparan Sulfate Sulfotransferases." The defense will be held at 2:00 p.m. in Kerr Hall, room 1001.

Abstract

Heparan sulfate (HS), a major component of the vascular system, is involved in regulating a number of functions of the blood vessel wall including blood coagulation, cell differentiation, and the inflammatory response. The wide range of biological functions makes HS an attractive target for the development of anticancer and anti-inflammatory agents.  Heparin, a specialized form of HS, is a commonly used anticoagulant drug. Clinical evidence has shown that cancer patients receiving antithrombotic agents had an increased survival rate (227), suggesting a unique therapeutic benefit of heparin.

Understanding the structure-function relationship of heparin will help improve its anticoagulant efficacy and allow us to take advantage of heparin’s antineoplastic activity. The long-term goal of our research involves using an enzyme-based approach to develop HS-based therapeutics for treating thrombotic diseases, cancer and excessive inflammatory responses.The biosynthesis of HS involves multiple specialized sulfotransferases, such as 2-O-sulfotransferase (2OST) and 6-O-sulfotransferase (6OST), which are essential for preparing HS with activities in regulating vascular development and blood coagulation. The stringent substrate specificity of HS sulfotransferases controls the sulfation patterns of HS, permitting HS to exhibit a specific function; however, limited knowledge regarding the mechanism of these enzymes has hindered our ability to prepare functionally specific HS. We aim to understand the mechanism of action of these two enzymes in hopes of developing heparin/HS with improved anticoagulant efficacy.

In this dissertation, we present successful crystallization of 2OST in complex with 3’-phosphoadenosine 5’-phosphate (PAP). The substrate recognition mechanism of 2OST was examined by way of extensive structurally guided mutational analysis. Several residues were identified including Arg-189, Tyr-94, and His-106 that are responsible for dictating the substrate specificity of 2OST. Despite success with the crystallization of 2OST, the journey towards crystallization of 6OST has been an ongoing process. A promising expression construct has been identified for 6OST-1. Using a homology model of 6OST-3 with structurally known 3OST-3, several residues involved in substrate binding and a potential catalytic base were proposed.