title: Quantitative Characterization of E-selectin Interaction with Native CD44 and P-selectin Glycoprotein Ligand-1 (PSGL-1) Using a Real Time Immunoprecipitation-based Binding Assay.
abstract: Selectins (E-, P-, and L-selectins) interact with glycoprotein ligands to mediate the essential tethering/rolling step in cell transport and delivery that captures migrating cells from the circulating flow. In this work, we developed a real time immunoprecipitation assay on a surface plasmon resonance chip that captures native glycoforms of two well known E-selectin ligands (CD44/hematopoietic cell E-/L-selectin ligand and P-selectin glycoprotein ligand-1) from hematopoietic cell extracts. Here we present a comprehensive characterization of their binding to E-selectin. We show that both ligands bind recombinant monomeric E-selectin transiently with fast on- and fast off-rates, whereas they bind dimeric E-selectin with remarkably slow on- and off-rates. This binding requires the sialyl Lewis x sugar moiety to be placed on both O- and N-glycans, and its association, but not dissociation, is sensitive to the salt concentration. Our results suggest a mechanism through which monomeric selectins mediate initial fast on and fast off kinetics to help capture cells out of the circulating shear flow; subsequently, tight binding by dimeric/oligomeric selectins is enabled to significantly slow rolling.
authors.affiliation: From the Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia.
authors.forename: Dina B
authors.initials: DB
authors.lastname: AbuSamra
authors.affiliation: From the Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia.
authors.forename: Alia
authors.initials: A
authors.lastname: Al-Kilani
authors.affiliation: From the Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia.
authors.forename: Samir M
authors.initials: SM
authors.lastname: Hamdan
authors.affiliation: From the Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Sakashita
authors.affiliation: From the Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia.
authors.forename: Samah Z
authors.initials: SZ
authors.lastname: Gadhoum
authors.affiliation: From the Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology, Thuwal 23955, Saudi Arabia jasmeen.merzaban@kaust.edu.sa.
authors.forename: Jasmeen S
authors.initials: JS
authors.lastname: Merzaban
mesh_terms: D045744:Cell Line, Tumor; D002465:Cell Movement; D019040:E-Selectin / Q000378:metabolism*; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D047468:Immunoprecipitation; D008562:Membrane Glycoproteins / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps
chemical_list: D019040:E-Selectin; D018960:Hyaluronan Receptors; D008562:Membrane Glycoproteins; C084882:P-selectin ligand protein
keywords: CD44; PSGL-1; cell adhesion; cell migration; glycobiology; glycoprotein; selectin; selectin ligand; sialyl Lewis x; surface plasmon resonance (SPR)