EXPRESSION AND BINDING-ACTIVITY OF THE CARBOXYL-TERMINAL PORTION OF THE CORE PROTEIN OF PG-M, A LARGE CHONDROITIN SULFATE PROTEOGLYCAN

PG-M is a large chondroitin sulfate proteoglycan that has been shown t o be expressed in the prechondrogenic condensation area of the develop ing chick limb buds. We previously isolated cDNA clones encoding the c ore protein of PG-M (Shinomura, T., Nishida, Y., Ito, H., and Kimata, K. (1993) J. Biol. Chem. 268, 14461-14469). The amino acid sequence de duced from the cDNA analysis revealed the presence of two epidermal gr owth factor-like domains, a C-type lectin-like domain, and a complemen t regulatory protein (CRP)-like domain at the COOH terminus. The COOH- terminal portion has been expressed as a fusion protein with glutathio ne S-transferase in Escherichia coli to test its carbohydrate binding activity using affinity chromatography. The purified fusion protein bi nds to immobilized D-mannose, D-galactose, L-fucose, and N-acetyl-D-gl ucosamine in a calcium-dependent manner. Furthermore, the fusion prote in binds to heparin- or heparan sulfate-Sepharose. To investigate role s of each COOH-terminal domain, we have made a truncated construct whi ch lacks the CRP-like domain and determined if the CRP-like domain is involved in the binding activity. The removal of this domain resulted in the complete loss of both C-type lectin-like and heparin binding ac tivities. The results suggest that a whole set of epidermal growth fac tor-, lectin-, and CRP-like domains may serve a functional structure f or these bindings.