ROTATIONAL MOBILITY OF THE FIBRINOGEN RECEPTOR GLYCOPROTEIN-IIB IIIA OR INTEGRIN ALPHA(IIB)BETA-3 IN THE PLASMA-MEMBRANE OF HUMAN PLATELETS/

Integrin alpha(IIb)beta3 or glycoprotein IIb/IIIa (GPIIb/IIIa, 228 kDa ) is a Ca2+-dependent, noncovalent heterodimer of glycoproteins IIb (G PIIb or alpha(IIb), 136 kDa) and IIIa (GPIIIa or beta3, 92 kDa), which serves as the receptor for fibrinogen and other adhesive proteins at the surface of activated platelets. We have determined the microsecond -range rotational motions of alpha(IIb)beta3 in resting platelets, in isolated plasma membranes, and reconstituted in 1-palmitoyl-2-oleoylph osphatidylcholine (POPC) bilayers. The measurements were based on the time-resolved phosphorescence anisotropy [r(t)] of erythrosin-labeled F(ab) fragments [Er-F(ab)] of monoclonal antibodies bound to alpha(IIb )beta3. In general, the r(t) decays were satisfactorily fitted to the sum of the two exponential terms and a constant, from which the initia l anisotropy (r(in) almost-equal-to 0.05-0.11), the short (phi1 almost -equal-to 1-14 mus) and the long (phi2 almost-equal-to 15-60 mus) rota tional correlation times, and the limiting anisotropy (r(infinity) alm ost-equal-to 0.02-0.07) were obtained. The observed values depended on the platelet preparation, temperature, Ca2+ concentration, and the an tibody used. In accordance with data on the order parameter and the vi scosity of the lipid bilayer of the platelet plasma membrane, phi2 and r(infinity) of the alpha(IIb)beta3-Er-F(ab) complexes in the three pr eparations decreased with the increase of temperature, the r(t) curves being fully reversible within the interval from 5 to 35-degrees-C. On the basis of direct and indirect evidence, we exclude both alpha(IIb) beta3 size heterogeneity, due to autoassociation or heteroassociation with membrane or cytoskeletal proteins, and the heterogeneous distribu tion of alpha(IIb)beta3 between lipid environments differing in microv iscosity as the source of the two correlation times. We interpret that phi1 and phi2 represent the segmental motion and global rotational re laxation of the alpha(IIb)beta3 monomer. The values of phi2, estimated for the two transmembrane helices of the alpha(IIb)beta3 monomer, are much lower than the experimental values, which suggests that interact ions between the lipid bilayer and ecto and/or endo domains of alpha(I Ib)beta3 further limit the rotational mobility of alpha(IIb)beta3. Fin ally, prolonged calcium chelation at 35-degrees-C, but not at 4-degree s-C, immobilized the majority of alpha(IIb)beta3 in the membrane, in a greement with previous functional and biochemical studies in whole pla telets and in isolated alpha(IIb)beta3 in solution.