RESEARCH RESOURCE FOR INTEGRATED GLYCOTECHNOLOGY:
Geert-Jan Boons

Development of Synthetic Carbohydrate Macro- and Micro-Array Technologies for Ligand Identification

Specific complex sulfated saccharide motifs in heparan sulfate (HS) confer selective binding and regulatory properties for many proteins. Detailed analysis of structure-activity relationships is beginning to unravel the biological significance of HS structural diversity. However, progress is hampered by the difficulties of identifying HS-binding motifs for specific proteins. It is to be expected that this problem can be addressed by screening a relatively large panel of well-defined HS fragments. Organic synthesis provides a powerful approach for obtaining well-defined HS fragments; however, no efficient strategy for the preparation of a wide range of HS structures has been reported. In order to address this issue, we will develop a modular approach for the chemical synthesis of a wide range of HS oligosaccharides. This modular approach will employ a set of properly protected disaccharide building blocks that resemble the 20 different disaccharide motifs found in HS. These disaccharides can then easily be assembled in a parallel combinatorial manner into larger structures. The disaccharide building blocks will be prepared from six strategically chosen monosaccharides, which are protected by a versatile set of orthogonal protecting groups (Lev, Fmoc, TBDPS and allyl). The fact that the monosaccharides and disaccharides will repeatedly be used will reduce the synthetic work considerably. Another key feature of the synthetic approach will be that at the end of the synthetic sequence, C-6 hydroxyls of glucosides and idosides will be selectively oxidized to carboxylic acids. This approach will avoid synthetic problems associated with iduronic and glucuronic acids. The resulting synthetic HS fragments will be employed for the development of macro- and micro-arrays and several approaches will be investigated for immobilization of the saccharides to a solid surface. The resulting arrays will be used to determine ligand requirements of a number of HS-binding proteins and anti-HS antibodies. In a second project, a library of N-acetyllactosamine derivatives will be synthesized using a mix and split approach, and innovative approaches will be developed for the discovery of high affinity carbohydrate-based ligands. This project may lead to compounds that are potent inhibitors of galectins and glycoprotein biosynthesis.