A microcantilever device to assess the effect of force on the lifetime of selectin-carbohydrate bonds.
A microcantilever technique was used to apply force to receptor-ligand molecules involved in leukocyte rolling on blood vessel walls. E-selectin was adsorbed onto 3-microm-diameter, 4-mm-long glass fibers, and the selectin ligand, sialyl Lewis(x), was coupled to latex microspheres. After binding, the microsphere and bound fiber were retracted using a computerized loading protocol that combines hydrodynamic and Hookean forces on the fiber to produce a range of force loading rates (force/time), r(f). From the distribution of forces at failure, the average force was determined and plotted as a function of ln r(f). The slope and intercept of the plot yield the unstressed reverse reaction rate, k(r)(o), and a parameter that describes the force dependence of reverse reaction rates, r(o). The ligand was titrated so adhesion occurred in approximately 30% of tests, implying that >80% of adhesive events involve single bonds. Monte Carlo simulations show that this level of multiple bonding has little effect on parameter estimation. The estimates are r(o) = 0.048 and 0.016 nm and k(r)(o) = 0.72 and 2.2 s(-1) for loading rates in the ranges 200-1000 and 1000-5000 pN s(-1), respectively. Levenberg-Marquardt fitting across all values of r(f) gives r(o) = 0.034 nm and k(r)(o) = 0.82 s(-1). The values of these parameters are in the range required for rolling, as suggested by adhesive dynamics simulations.
Tees
D F
DF
Department of Chemical Engineering and Institute of Medicine and Engineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Waugh
R E
RE
Hammer
D A
DA
D000268:Adhesiveness; D055592:Biophysical Phenomena; D001703:Biophysics; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry*; D004151:Dipeptides; D019040:E-Selectin / Q000737:chemistry / Q000378:metabolism; D004867:Equipment Design; D066298:In Vitro Techniques; D007700:Kinetics; D007840:Latex; D008024:Ligands; D008863:Microspheres; D008954:Models, Biological; D009010:Monte Carlo Method; D009844:Oligosaccharides; D011485:Protein Binding; D019087:Selectins / Q000737:chemistry* / Q000378:metabolism*; D000080506:Sialyl Lewis X Antigen; D014743:Videotape Recording
C415363:1, 3-di-(N-carboxybenzoyl-leucyl-leucyl)amino acetone; D002241:Carbohydrates; D004151:Dipeptides; D019040:E-Selectin; D007840:Latex; D008024:Ligands; D009844:Oligosaccharides; D019087:Selectins; D000080506:Sialyl Lewis X Antigen