Changes in glycoconjugate structure are associated with a wide range of normal physiological and developmental processes as well as with disease conditions such as oncogenic transformation, suggesting that carbohydrates play an important role in these biological phenomena. However, the ability of oligosaccharide portion of biological glycoconjugates to carry information depends on the existence of recognition mechanisms for these complex carbohydrate structures. Recent studies have revealed the existence of a family of calcium ion-dependent carbohydrate recognition domains (CRDs) which mediate selective binding to saccharide structure by proteins with diverse biological functions. Proteins containing such """"""""C-type"""""""" CRDs range from serum mannose- binding proteins and hepatic membrane receptors for endocytosis of asialoglycoproteins, to pulmonary surfactant apoprotein and core protein of cartilage proteoglycan. It thus appears that this class of domains represents a widespread mechanism for decoding information in complex carbohydrates. The goal of the proposed work is to establish the molecular basis for the selective binding of C-type CRDs to specific oligosaccharides. Expression systems lore producing the various C-type CRDs in Escherichia coli are being developed. The ability to produce isolated, homogeneous CRDs will form the basis for a detailed investigation of CRD structure and function. Experimental approaches will include the following: (1) studies on the mechanism of carbohydrate Ligation using a combination of mutagenesis and physical methods, including crystallography; (2) analysis of conformational changes associated with calcium ion- dependence of ligand-binding by the domains; (3) definition of binding specificity and endogenous ligands for proteins containing C-type CRDs; and (4) demonstration of saccharide-binding activity in additional proteins predicted (from primary structure analysis) to have CRDs. Studies designed to establish evolutionary relationships within the family of C-type CRDs are also proposed.
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