Time and force dependence of the rupture of glycoprotein IIb-IIIa-fibrinogen bonds between latex spheres

Citation
Hl. Goldsmith et al., Time and force dependence of the rupture of glycoprotein IIb-IIIa-fibrinogen bonds between latex spheres, BIOPHYS J, 78(3), 2000, pp. 1195-1206
Citations number
56
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOPHYSICAL JOURNAL
ISSN journal
00063495 → ACNP
Volume
78
Issue
3
Year of publication
2000
Pages
1195 - 1206
Database
ISI
SICI code
0006-3495(200003)78:3<1195:TAFDOT>2.0.ZU;2-5
Abstract
We studied the shear-induced breakup of doublets of aldehyde/sulfate (A/S) latex spheres covalently linked with purified:platelet GPIIb-IIIa receptor, and cross-linked by fibrinogen. Flow cytometry with fluorescein isothiocya nate-fibrinogen showed than an average of 22,500 molecules of active GPIIb- IIIa were captured per sphere, with a mean K-d = 56 nM for fibrinogen bindi ng. The spheres, suspended in buffered 19% Ficoll 400 containing 120 or 240 pM fibrinogen, were subjected to Couette flow in a counter-rotating cone-p late rheoscope. Doublets, formed by two-body collisions at low shear rate ( G = 8 s(-1)) for less than or equal to 15 min, were subjected to shear stre ss from 0.6 to 2.9 Nm(-2), their rotations recorded until they broke up or were lost to view. Although breakup was time dependent, occurring mostly in the first 2 rotations after the onset of shear, the percentage of doublets broken up after 10 rotations were almost independent of normal hydrodynami c force, F-n: at 240 pN, 15.6, 16.0, and 17.0% broke up in the force range 70-150 pN, 150-230 pN, and 230-310 pN, Unexpectedly, at both [fibrinogen], the initial rate of breakup was highest in the lowest force range, and comp uter simulation using a stochastic model of breakup was unable to simulate the time course of breakup. When pre-sheared at low G for >15 min, no doubl ets broke up within 10 rotations at 70 < F-n < 310 pN; it required >3 min s hear (>1110 rotations) at F-n = 210 pN for significant breakup to occur. Ot her published work has shown that binding of fibrinogen to GPIIb-IIIa immob ilized on plane surfaces exhibits an initial fast reversible process with r elative low affinity succeeded by transformation of GPIIb-IIIa to a stable high-affinity complex. We postulate that most doublet breakups observed wit hin 10 rotations were from a population of young doublets having low number s of bonds, by dissociation of the initial receptor complex relatively unre sponsive to force. The remaining, older doublets with GPIIb-IIIa in the hig h-affinity complex were not broken up in the time or range of forces studie d.