Expression of isolated C-type carbohydrate recognition domains.
A galactose-binding lectin from the venom of the snake Trimeresurus stejnegeri consists of isolated carbohydrate recognition domains, belonging to group VII of the C-type animal lectins. As a first step toward determining the tertiary structure of the galactose-specific lectin, we produced the lectin in Escherichia coli. By in vitro refolding and affinity chromatography, modest amounts (8 mg/liter) of active recombinant proteins were obtained. The recombinant protein was homogeneous, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and mass spectrometry. Its amino acid sequence without the initiated methionine at the N-terminus was also characterized by mass spectrometry. The data of hemagglutination and enzyme-linked lectin binding assays demonstrated that the recombinant lectin showed similar sugar-binding activity as the native protein. In addition, fluorescence spectroscopy and circular dichroism also showed obviously their structural similarity.
Xu
Q
Q
Shanghai Institute of Biochemistry, Chinese Academy of Sciences, 320 Yue-Yang Road, Shanghai, 200031, People's Republic of China.
Zeng
R
R
Wu
X F
XF
Wang
K Y
KY
D000818:Animals; D001667:Binding, Competitive; D002942:Circular Dichroism; D003435:Crotalid Venoms / Q000737:chemistry*; D004926:Escherichia coli; D005697:Galactosides / Q000378:metabolism; D037161:Galectins; D006384:Hemagglutination; D006388:Hemagglutinins / Q000737:chemistry* / Q000235:genetics; D006601:Hexoses / Q000494:pharmacology; D013058:Mass Spectrometry; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D017841:Trimeresurus
D003435:Crotalid Venoms; D005697:Galactosides; D037161:Galectins; D006388:Hemagglutinins; D006601:Hexoses; D011994:Recombinant Proteins