title: Calcium cation regulation of glycoprotein IIb-IIIa complex formation in platelet plasma membranes.
abstract: The regulating effect of Ca2+ on the association and dissociation of the glycoprotein IIb-IIIa complex from human platelet membranes was determined both for detergent-solubilized and intact plasma membranes. Glycoproteins IIb and IIIa were solubilized from isolated membranes with 0.5% Triton X-100 and incubated in buffers containing ionized calcium, which resulted in the formation of the glycoprotein IIb-IIIa complex. With the addition of EGTA to reduce the ionized calcium content of the solution, the glycoprotein IIb-IIIa complex dissociated. This dissociation was measured by comparing the sedimentation properties of the glycoproteins and by observing the susceptibility of glycoprotein IIb to thrombin-catalyzed hydrolysis. With 10(-3) M Ca2+, glycoproteins IIb and IIIa were resistant to hydrolysis at thrombin concentrations up to 2.4 X 10(-5) M. When the Ca2+ concentration was decreased to less than 10(-4) M by chelation with EDTA or EGTA, glycoprotein IIb was cleaved by thrombin. This increased susceptibility to thrombin hydrolysis at decreasing Ca2+ levels correlated with the increased dissociation of the glycoprotein IIb-IIIa complex as determined by sucrose density centrifugation. Susceptibility to thrombin hydrolysis was also used as a probe to determine the extent to which Ca2+ regulates the formation of the glycoprotein IIb-IIIa complex within membranes. At more than micromolar levels of Ca2+, less than 10% of the membrane-bound glycoprotein IIb was cleaved by thrombin. Increased hydrolysis was observed at decreasing concentrations of Ca2+. Resistance to thrombin hydrolysis was partially regained upon the readdition of Ca2+ to dissociated glycoproteins. These data indicate that micromolar concentrations of Ca2+ exert a direct effect on platelet plasma membrane structure by regulating the intramembranous interactions of glycoprotein IIb.
mesh_terms: D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D006023:Glycoproteins / Q000097:blood*; D006801:Humans; D006868:Hydrolysis; D017830:Octoxynol; D010980:Platelet Membrane Glycoproteins; D011092:Polyethylene Glycols; D013917:Thrombin / Q000494:pharmacology
chemical_list: D006023:Glycoproteins; D010980:Platelet Membrane Glycoproteins; D011092:Polyethylene Glycols; D017830:Octoxynol; D013917:Thrombin; D002118:Calcium