title: Leukocyte function-associated antigen 1-mediated adhesion stability is dynamically regulated through affinity and valency during bond formation with intercellular adhesion molecule-1.
abstract: Neutrophil rolling and transition to arrest on inflamed endothelium are dynamically regulated by the affinity of the beta(2) integrin CD11a/CD18 (leukocyte function associated antigen 1 (LFA-1)) for binding intercellular adhesion molecule (ICAM)-1. Conformational shifts are thought to regulate molecular affinity and adhesion stability. Also critical to adhesion efficiency is membrane redistribution of active LFA-1 into dense submicron clusters where multimeric interactions occur. We examined the influences of affinity and dimerization of LFA-1 on LFA-1/ICAM-1 binding by engineering a cell-free model in which two recombinant LFA-1 heterodimers are bound to respective Fab domains of an antibody attached to latex microspheres. Binding of monomeric and dimeric ICAM-1 to dimeric LFA-1 was measured in real time by fluorescence flow cytometry. ICAM-1 dissociation kinetics were measured while LFA-1 affinity was dynamically shifted by the addition of allosteric small molecules. High affinity LFA-1 dissociated 10-fold faster when bound to monomeric compared with dimeric ICAM-1, corresponding to bond lifetimes of 25 and 330 s, respectively. Downshifting LFA-1 into an intermediate affinity state with the small molecule I domain allosteric inhibitor IC487475 decreased the difference in dissociation rates between monomeric and dimeric ICAM-1 to 4-fold. When LFA-1 was shifted into the low affinity state by lovastatin, both monomeric and dimeric ICAM-1 dissociated in less than 1 s, and the dissociation rates were within 50% of each other. These data reveal the respective importance of LFA-1 affinity and proximity in tuning bond lifetime with ICAM-1 and demonstrate a nonlinear increase in the bond lifetime of the dimer versus the monomer at higher affinity.
authors.affiliation: Department of Biomedical Engineering, University of California, Davis, California 95616, USA.
authors.forename: Melissa R
authors.initials: MR
authors.lastname: Sarantos
authors.forename: Subhadip
authors.initials: S
authors.lastname: Raychaudhuri
authors.forename: Aaron F H
authors.initials: AF
authors.lastname: Lum
authors.forename: Donald E
authors.initials: DE
authors.lastname: Staunton
authors.forename: Scott I
authors.initials: SI
authors.lastname: Simon
mesh_terms: D000494:Allosteric Regulation; D002448:Cell Adhesion / Q000502:physiology*; D002474:Cell-Free System; D019281:Dimerization; D004730:Endothelium, Vascular / Q000502:physiology; D005434:Flow Cytometry; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism / Q000502:physiology*; D007700:Kinetics; D016169:Lymphocyte Function-Associated Antigen-1 / Q000378:metabolism / Q000502:physiology*; D008954:Models, Biological; D009504:Neutrophils / Q000502:physiology*; D011487:Protein Conformation
chemical_list: D016169:Lymphocyte Function-Associated Antigen-1; D018799:Intercellular Adhesion Molecule-1