FROM GENE TO STRUCTURE: Formation of Iduronic Acid in Dermatan Sulfate by Two DS-epimerases

Abstract: During embryonic development and adult life a wide range of cell behaviors such as differentiation, proliferation and migration are in effect to maintain tissue integrity and function. An integral part of these dynamic processes is the interplay between the cells and their environment, i.e. the extracellular space. Complex polysaccharides, such as dermatan sulfate play a key role in these processes. Dermatan sulfate is a long linear polysaccharide of a repeating disaccharide unit consisting of N-acetylgalactosamine and glucuronic acid/iduronic acid. Formation of iduronic acid through epimerisation of glucuronic acid is a key modification in dermatan sulfate biosynthesis. In the present thesis two enzymes have been identified that catalyzes this reaction, i.e. DS-epimerase 1 and 2. By using fold-recognition modeling a structural model was obtained for DS-epimerase 1, which enabled characterization of the active site of the enzyme with mutational analysis. A feature of a dermatan sulfate chain is the presence of sequences of iduronic acid containing disaccharides separated by stretches of glucuronic acid containing disaccharides. The former are also commonly associated with sulfation in the 4-O-position of N-acetylgalactosamine suggesting that there is interplay between sulfation and epimerization. In this thesis it is shown that both epimerases are simultaneously needed together with the sulfotransferase D4ST-1 to generate these types of domains. Interestingly, 4-O-sulfated iduronic acid blocks constitute the binding site for several growth factors, including fibroblast growth factor 2. Through this binding, dermatan sulfate can sequester the growth factor in the extracellular matrix and regulate the availability of the growth factor at the cell surface. Dynamic changes in dermatan sulfate structure thus offers a way to fine-tune signaling. In this thesis evidence is presented that common growth factors such as transforming growth factor-?, platelet derived growth factor and epidermal growth factor have the ability to regulate both epimerase activity and gene expression of several sulfotransferases, including D4ST-1, which results in reduction of 4-O-sulfated iduronic acid blocks.