Mammalian Sialic Acid Binding Proteins

Scripps Research Institute, La Jolla CA

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Abstract

This project is directed to the characterization of the recently described siglec family of cell adhesion receptors, a subfamily of the immunoglobulin superfamily. The extracellular domains of the siglecs have a homologous N-terminal Ig domain which confers the ability to bind to sialic acid containing carbohydrate groups of glycoproteins and glycolipids. To date there are ten known members of the family, nine of which are expressed in cells of the immune system. Most is known about CD22 (siglec 2), which has been demonstrated to be an important regulatory molecule in the B cell antigen receptor complex, and has been implicated in dampening the immune response against autoantigens and autoimmune disease. In contrast to CD22, the detailed biology and functions of the other siglecs are poorly understood. The primary goal of the project will be to characterize the detailed specificities of murine and human siglecs towards sialyloligosaccharides of glycoproteins and glycolipids, and to develop multivalent sialyloligosaccharide probes unique to each siglec that will aid in their identification and characterization in complex biological systems. The initial objective will be to establish the affinity and specificity of each of the 11 known siglecs for >20 common sialyloligosaccharides found on carbohydrate groups of glycoproteins and glycolipids. The strategy will be to synthesize monovalent sialyloligosaccharides using a chemo-enzymatic approach, and establish the relative binding affinities using ELISA based and BiaCore based binding assays. This information will be used to design synthetic multivalent sialyloligosaccharide probes that distinguish each selectin in complex biological systems that will allow them to be used as tools in exploring their biological functions. A key biological resource to this project will be the availability of 'knockout mice' deficient in sialyltransferase genes involved in the synthesis of the sialoside ligands of siglecs. These mice will aid in the evaluation of the functional status of siglecs and provide insights into the role of these enzymes in the regulation of siglec function.