title: Galectins bind to the multivalent glycoprotein asialofetuin with enhanced affinities and a gradient of decreasing binding constants.
abstract: Our previous isothermal titration microcalorimetry (ITC) studies of the binding of synthetic multivalent carbohydrates to the Man/Glc-specific lectins concanavalin A (ConA) and Dioclea grandiflora lectin (DGL) showed negative binding cooperativity that was due to the carbohydrate ligands and not the proteins [Dam, T. K., et al. (2002) Biochemistry 41, 1351-1358]. The negative cooperativity was associated with the decreasing functional valence of the carbohydrates upon progressive binding of their epitopes. The present study also shows negative cooperativity in the ITC binding data of asialofetuin (ASF), a glycoprotein that possesses nine LacNAc epitopes, to galectin-1, -2, -3, -4, -5, and -7, and truncated, monomer versions of galectin-3 and -5, which are members of a family of animal lectins. Although the observed K(a) values for binding of ASF to the galectins and two truncated forms are only 50-80-fold greater than that of LacNAc, analysis of the data in terms of the relationship between the observed macroscopic free energy of binding and the decreasing microscopic free energies of binding of the epitopes shows that the first LacNAc epitope of ASF binds with approximately 6000-fold higher affinity than the last epitope. Thus, the microscopic binding constants of the galectins for the first epitope(s) of ASF are in the nanomolar range, with a gradient of decreasing binding constants of the remaining epitopes. The results indicate that the above galectins bind with fractional, high affinities to multivalent glycoproteins such as ASF, independent of the quaternary structures of the galectins. These findings have important implications for the binding of galectins to multivalent carbohydrate receptors.
authors.affiliation: Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors.forename: Tarun K
authors.initials: TK
authors.lastname: Dam
authors.forename: Hans-J
authors.initials: HJ
authors.lastname: Gabius
authors.forename: Sabine
authors.initials: S
authors.lastname: André
authors.forename: Herbert
authors.initials: H
authors.lastname: Kaltner
authors.forename: Martin
authors.initials: M
authors.lastname: Lensch
authors.forename: C Fred
authors.initials: CF
authors.lastname: Brewer
mesh_terms: D000818:Animals; D001212:Asialoglycoproteins / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002417:Cattle; D003001:Cloning, Molecular; D017931:DNA Primers; D060748:Fetuins; D037502:Galectin 3 / Q000737:chemistry / Q000378:metabolism; D037541:Galectin 4 / Q000737:chemistry / Q000378:metabolism; D037161:Galectins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D011485:Protein Binding; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D000509:alpha-Fetoproteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D001212:Asialoglycoproteins; D017931:DNA Primers; D060748:Fetuins; D037502:Galectin 3; D037541:Galectin 4; D037161:Galectins; D011994:Recombinant Proteins; D000509:alpha-Fetoproteins; C017029:asialofetuin