SBIR-STTR Award

Access to a library of structurally defined N-glycans is an urgent scientific need for glyco-arrays for rapid analysis of carbohydrate binding proteins (CBPs), development of diagnostics and therapeutics for many diseases. However, only a small number of N-glycans are currently commercially available, which hampered researches in Glycobiology. This application will chemo-enzymatically prepare a library of 80 N-glycans (hybrid-, linear and complex-type) and commercialize them. The long-term goal is to produce thousands of natural N-glycans for carbohydrate-based highthroughput assays. Firstly, essential N-glycan precursors will be prepared in gram scales via either chemical synthesis or isolation from chicken eggs yolk and white. These N-glycans will then be enzymatically elongated or digested to yield 80 different N-glycans with defined structures (1 mg), among which only 12 N-glycans are currently available from commercial sources. The N-glycans contain 5 to 11 monosaccharide residues. This process involves several well characterized glycosyltransferases, glycoside hydrolases and other enzymes. All the N-glycans will be purified to 98%, and fully characterized by NMR and MS.

Project Terms:
Anions; base; Biological Assay; carbohydrate binding protein; Carbohydrates; chemical synthesis; Chickens; Chromatography; Complex; Custom; detector; Development; Diagnostic; Disease; egg; Egg Yolk; Ensure; Enzymes; Generations; Glycobiology; Glycoside Hydrolases; glycosyltransferase; Goals; High Pressure Liquid Chromatography; Hybrids; Libraries; meetings; Monosaccharides; Oligosaccharides; Ovalbumin; Performance; Physiologic pulse; Polysaccharides; Process; Reference Standards; Research; Small Business Innovation Research Grant; Source; Specificity; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Structure; Testing; Therapeutic

Access to structurally defined N-glycans is an urgent scientific need for glycan arrays for rapid analysis of carbohydrate binding proteins (CBPs), development of diagnostics and therapeutics for many diseases, as well as glycoanalysis. However, only a small number of N-glycans are currently commercially available, most of which are symmetric ones. The lack of structurally defined glycans has greatly hampered research in Glycobiology. Due to complexity and low abundance of N-glycans, it is extremely difficult to separate and purify homogenous N-glycans from natural resources. Thus chemical synthesis of N-glycans has been pursued instead. However, very limited N-glycans have been synthesized due to challenges in the chemical methodology. In the preceding Phase I proposal, the contractor developed a facile and efficient chemical core synthesis with an enzymatic extension (CCSEE) strategy and a rapid HPLC-based purification approach for N-glycan synthesis. The contractor successfully prepared 75 structurally defined N-glycans with a purity of 98%. In this Phase II project, the Contractor shall further improve the CCSEE strategy for synthesis of cancer-related N-glycans, including Neu5Gc-terminated, core-fucosylated, bisected, and 1,6 branched N-glycans. These glycans cover the chemical space of bi-, tri- and tetra-antennary complex-type N-glycans. This process involves several well-characterized glycosyltransferases, glycoside hydrolases, and other enzymes. All the proposed N-glycans shall be purified to 98%, and fully characterized by HPLC, MS, and partially by NMR.