title: Rolling adhesion of alphaL I domain mutants decorrelated from binding affinity.
abstract: Activated lymphocyte function-associated antigen-1 (LFA-1, alphaLbeta2 integrin) found on leukocytes facilitates firm adhesion to endothelial cell layers by binding to intercellular adhesion molecule-1 (ICAM-1), which is up-regulated on endothelial cells at sites of inflammation. Recent work has shown that LFA-1 in a pre-activation, low-affinity state may also be involved in the initial tethering and rolling phase of the adhesion cascade. The inserted (I) domain of LFA-1 contains the ligand-binding epitope of the molecule, and a conformational change in this region during activation increases ligand affinity. We have displayed wild-type I domain on the surface of yeast and validated expression using I domain specific antibodies and flow cytometry. Surface display of I domain supports yeast rolling on ICAM-1-coated surfaces under shear flow. Expression of a locked open, high-affinity I domain mutant supports firm adhesion of yeast, while yeast displaying intermediate-affinity I domain mutants exhibit a range of rolling phenotypes. We find that rolling behavior for these mutants fails to correlate with ligand binding affinity. These results indicate that unstressed binding affinity is not the only molecular property that determines adhesive behavior under shear flow.
authors.affiliation: Department of Bioengineering, University of Pennsylvania, 220 South 33rd Street, Philadelphia, PA 19104, USA.
authors.forename: Lauren R
authors.initials: LR
authors.lastname: Pepper
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Hammer
authors.forename: Eric T
authors.initials: ET
authors.lastname: Boder
mesh_terms: D040881:CD11a Antigen / Q000737:chemistry*; D000939:Epitopes / Q000737:chemistry; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000737:chemistry*; D007700:Kinetics; D016169:Lymphocyte Function-Associated Antigen-1 / Q000737:chemistry*; D008968:Molecular Conformation; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry; D000267:Tissue Adhesions; D015854:Up-Regulation
chemical_list: D040881:CD11a Antigen; D000939:Epitopes; D016169:Lymphocyte Function-Associated Antigen-1; D029701:Saccharomyces cerevisiae Proteins; D018799:Intercellular Adhesion Molecule-1