title: Consensus repeat domains of E-selectin enhance ligand binding.
abstract: To study the structural characteristics of E-selectin necessary for mediating cell adhesion, we examined the role of the consensus repeat (CR) domains in E-selectin function. Soluble constructs containing different numbers of CR domains were stably expressed in Chinese hamster ovary cells, purified to homogeneity, and characterized. The minimum functional unit of soluble E-selectin consisted of the lectin (Lec) and epidermal growth factor (EGF) domains alone (Lec-EGF) as indicated by its ability to mediate in vitro HL-60 cell adhesion. However, E-selectin containing all six CR domains (Lec-EGF-CR6) at its COOH terminus was the most potent in blocking neutrophil or HL-60 cell adhesion to either immobilized E-selectin or cytokine-stimulated human umbilical vein endothelial cells. This increased potency of Lec-EGF-CR6 in blocking cell adhesion was not due to CR-mediated oligomerization of the protein. Lec-EGF-CR6 was most likely monomeric in solution, as judged by gel filtration fast protein liquid chromatography, membrane ultrafiltration, and chemical cross-linking analysis. Therefore, although the lectin and EGF domains are necessary and sufficient for mediating cell adhesion, the additional six CR domains, present in native E-selectin, contribute to the enhanced binding of E-selectin to its ligand.
authors.affiliation: Department of Inflammation/Autoimmune Diseases, Hoffmann-La Roche Inc., Nutley, New Jersey 07110.
authors.forename: S H
authors.initials: SH
authors.lastname: Li
authors.forename: D K
authors.initials: DK
authors.lastname: Burns
authors.forename: J M
authors.initials: JM
authors.lastname: Rumberger
authors.forename: D H
authors.initials: DH
authors.lastname: Presky
authors.forename: V L
authors.initials: VL
authors.lastname: Wilkinson
authors.forename: M
authors.initials: M
authors.lastname: Anostario
authors.forename: B A
authors.initials: BA
authors.lastname: Wolitzky
authors.forename: C R
authors.initials: CR
authors.lastname: Norton
authors.forename: P C
authors.initials: PC
authors.lastname: Familletti
authors.forename: K J
authors.initials: KJ
authors.lastname: Kim
mesh_terms: D001483:Base Sequence; D001667:Binding, Competitive; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000378:metabolism*; D016384:Consensus Sequence; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000737:chemistry; D019040:E-Selectin; D006801:Humans; D066298:In Vitro Techniques; D008024:Ligands; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins; D012091:Repetitive Sequences, Nucleic Acid; D017384:Sequence Deletion; D012995:Solubility; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
chemical_list: D015815:Cell Adhesion Molecules; D017931:DNA Primers; D019040:E-Selectin; D008024:Ligands; D011994:Recombinant Proteins