Abstract
A recombinant polypeptide composed of the alpha-helical neck region and carbohydrate recognition domain (CRD) of bovine conglutinin was expressed in Escherichia coli. The recombinant protein formed inclusion bodies but could be solubilised using a denaturation-renaturation cycle based on urea and then purified by affinity chromatography on a TSK-N-acetylglucosamide column. The purified product behaved as a homotrimer in non-dissociating conditions, with three CRDs held together by the alpha-helical neck regions. The trimer, although lacking the N-terminal and collagen regions of the native conglutinin, showed the same binding carbohydrate specificities as the native molecule, for the complement fragment C3b and for lipopolysaccharides derived from Gram-negative bacteria.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Base Sequence
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Binding, Competitive
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Calcium / metabolism
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Calcium / pharmacology
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Cattle
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Chromatography, Agarose
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Collectins*
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Complement C3b / metabolism
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Edetic Acid / metabolism
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Edetic Acid / pharmacology
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Electrophoresis, Polyacrylamide Gel
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Gene Expression Regulation, Bacterial
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Lectins / genetics
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Lectins / metabolism*
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Lipopolysaccharides / metabolism
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Molecular Sequence Data
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Monosaccharides / metabolism
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Monosaccharides / pharmacology
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Peptides / chemistry*
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Peptides / genetics
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Peptides / metabolism*
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Protein Binding / genetics
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Protein Structure, Secondary
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Recombinant Proteins / chemistry
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Recombinant Proteins / isolation & purification
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Recombinant Proteins / metabolism
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Serum Globulins / genetics
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Serum Globulins / metabolism*
Substances
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Collectins
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Lectins
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Lipopolysaccharides
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Monosaccharides
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Peptides
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Recombinant Proteins
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Serum Globulins
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conglutinin
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Complement C3b
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Edetic Acid
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Calcium