title: Biochemical and biophysical characterization of human recombinant IgE-binding protein, an S-type animal lectin.
abstract: IgE-binding protein (epsilon BP) was originally identified by virtue of its affinity for IgE. It is now known to be a beta-galactoside-binding lectin with the characteristic of an S-type carbohydrate recognition domain. The protein is composed of two domains: the amino-terminal domain consisting of tandem repeats and the carboxyl-terminal domain containing sequences shared by other S-type carbohydrate recognition domains. The amino-terminal domain also contains a number of potential recognition sites for collagenase cleavage. In this study, human epsilon BP was first expressed in Escherichia coli, and the carboxyl-terminal domain (epsilon BP-C) was then generated by collagenase digestion of epsilon BP. By equilibrium dialysis, the association constants of epsilon BP and epsilon BP-C for lactose were found to be similar (6.0 +/- 0.70) x 10(4) M-1 and (4.7 +/- 0.27) x 10(4) M-1, respectively. Both polypeptides contain only one lactose-binding site/molecule. By an assay involving binding of 125I-labeled epsilon BP or epsilon BP-C to solid phase IgE, and inhibition of this binding by saccharides, it was determined that epsilon BP-C retains the saccharide specificity of epsilon BP. Importantly, although unlabeled epsilon BP-C inhibited the binding of the radiolabeled epsilon BP to IgE, unlabeled epsilon BP caused increased binding to IgE, suggesting self-association among epsilon BP molecules. Oligomeric structures resulting from self-association of epsilon BP were confirmed by chemical cross-linking studies. Furthermore, epsilon BP possesses hemagglutination activity on rabbit erythrocytes, whereas epsilon BP-C lacks such activity. Based on these results, we propose a structural model for multivalency of epsilon BP: dimerization or oligomerization of epsilon BP occurs through intermolecular interaction involving the amino-terminal domain.
authors.affiliation: Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037.
authors.forename: D K
authors.initials: DK
authors.lastname: Hsu
authors.forename: R I
authors.initials: RI
authors.lastname: Zuberi
authors.forename: F T
authors.initials: FT
authors.lastname: Liu
mesh_terms: D000818:Animals; D000943:Antigens, Differentiation / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D002241:Carbohydrates / Q000494:pharmacology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D037502:Galectin 3; D005822:Genetic Vectors; D006384:Hemagglutination; D006801:Humans; D007700:Kinetics; D007785:Lactose / Q000494:pharmacology; D037102:Lectins / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011487:Protein Conformation; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
chemical_list: D000943:Antigens, Differentiation; D002241:Carbohydrates; D037502:Galectin 3; D037102:Lectins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D007785:Lactose