Ligand affinity of the 67-kD elastin/laminin binding protein is modulated by the protein's lectin domain: visualization of elastin/laminin-receptor complexes with gold-tagged ligands.
Video-enhanced microscopy was used to examine the interaction of elastin- or laminin-coated gold particles with elastin binding proteins on the surface of live cells. By visualizing the binding events in real time, it was possible to determine the specificity and avidity of ligand binding as well as to analyze the motion of the receptor-ligand complex in the plane of the plasma membrane. Although it was difficult to interpret the rates of binding and release rigorously because of the possibility for multiple interactions between particles and the cell surface, relative changes in binding have revealed important aspects of the regulation of affinity of ligand-receptor interaction in situ. Both elastin and laminin were found to compete for binding to the cell surface and lactose dramatically decreased the affinity of the receptor(s) for both elastin and laminin. These findings were supported by in vitro studies of the detergent-solubilized receptor. Further, immobilization of the ligand-receptor complexes through binding to the cytoskeleton dramatically decreased the ability of bound particles to leave the receptor. The changes in the kinetics of ligand-coated gold binding to living cells suggest that both laminin and elastin binding is inhibited by lactose and that attachment of receptor to the cytoskeleton increases its affinity for the ligand.
Mecham
R P
RP
Department of Medicine, Washington University Medical School, St. Louis, Missouri 63110.
Whitehouse
L
L
Hay
M
M
Hinek
A
A
Sheetz
M P
MP
D008841:Actin Cytoskeleton / Q000502:physiology; D000818:Animals; D001667:Binding, Competitive; D001692:Biological Transport; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D004058:Diffusion; D004549:Elastin / Q000378:metabolism*; D005697:Galactosides / Q000378:metabolism; D007797:Laminin / Q000378:metabolism*; D037102:Lectins; D008024:Ligands; D008560:Membrane Fluidity; D008854:Microscopy, Electron; D008970:Molecular Weight; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism*; D017474:Receptors, Laminin; D014741:Video Recording
D005697:Galactosides; D007797:Laminin; D037102:Lectins; D008024:Ligands; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D017474:Receptors, Laminin; C054901:elastin-binding proteins; D004549:Elastin