Expression of isolated C-type carbohydrate recognition domains

A galactose-binding lectin from the venom of the snake Trimeresurus stejneg eri consists of isolated carbohydrate recognition domains, belonging to gro up VII of the C-type animal lectins. As a first step toward determining the tertiary structure of the galactose-specific lectin, we produced the lecti n in Escherichia coil. By in vibro refolding and affinity chromatography, m odest amounts (8 mg/liter) of active recombinant proteins were obtained. Th e recombinant protein was homogeneous, as determined by sodium dodecyl sulf ate-polyacrylamide gel electrophoresis and mass spectrometry. Its amino aci d sequence without the initiated methionine at the N-terminus was also char acterized by mass spectrometry. The data of hemagglutination and enzyme-lin ked lectin binding assays demonstrated that the recombinant lectin showed s imilar sugar-binding activity as the native protein. In addition, fluoresce nce spectroscopy and circular dichroism also showed obviously their structu ral similarity. (C) 2000 Academic Press.