We have recently shown that human monocytes and U937 cells possess two molecular classes of Fc gamma receptor. One, a 72,000-mol-wt sialoglycoprotein, has high affinity for certain subclasses of human and murine monomeric IgG. The other is a 40,000-mol-wt protein (p40) with low affinity for monomeric IgG but with the capacity to bind IgG aggregates or IgG-coated particles. In the present study, a 40,000-mol-wt single chain protein, apparently identical to p40 from U937 cells, was isolated from surface-radioiodinated human platelets by affinity purification using a murine IgG2b monoclonal antibody to p40. This 40,000-mol-wt protein was not seen when control IgG2b or unrelated murine monoclonal antibodies were employed in place of anti-p40. The same 40,000-mol-wt protein was also recovered from an IgG-Sepharose affinity adsorbent, but not from ovalbumin-or myoglobin-Sepharose. The 72,000-mol-wt Fc gamma receptor of monocytes was not identified on platelets. Monoclonal anti-p40 and Fab fragments derived from this antibody blocked platelet aggregation by heat-aggregated human IgG, whereas a control murine IgG2b protein had little or no inhibitory effect at 500-1,000-fold higher concentrations. A murine IgG1 monoclonal antibody, reactive with an unrelated platelet-specific membrane antigen, did not inhibit platelet responses to aggregated IgG. Anti-p40 did not affect platelet aggregation by thrombin, collagen, or fibrinogen plus ADP. Although anti-p40 did not directly aggregate platelets in the concentrations employed, cross-linking with F(ab')2 goat anti-murine Ig induced apyrase-sensitive aggregation of anti-p40-treated platelets. This indicates that p40 possesses transmembrane linkage for platelet activation and secretion. These observations strongly suggest that this newly recognized 40,000-mol-wt platelet membrane protein serves as an Fc gamma receptor.

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Human platelet Fc receptor for immunoglobulin G. Identification as a 40,000-molecular-weight membrane protein shared by monocytes.

Human platelet glycoprotein Ib (GP Ib) is a major integral membrane protein that has been identified as the platelet-binding site mediating the factor VIII/von Willebrand-factor-dependent adhesion of platelets to vascular subendothelium. Recent evidence suggests that GP Ib is normally complexed with another platelet membrane protein, GP IX. In this study, human platelet plasma membranes were selectively solubilized with a buffer containing 0.1% (v/v) Triton X-100. The GP Ib complex (GP Ib plus GP IX) was purified to homogeneity in ∼ 30% yield by immunoaffinity chromatography of the membrane extract using the anti-(glycoprotein Ib complex) murine monoclonal antibody, WM 23, coupled to agarose. GP Ib and GP IX were subsequently isolated as purified components by immunoaffinity chromatography of the GP Ib complex using a second anti-(glycoprotein Ib complex) monoclonal antibody, FMC 25, coupled to agarose. As assessed by dodecyl sulphate/polyacrylamide gel electrophoresis, purified GP Ib was identical to the molecule on intact platelets and had an apparent relative molecular mass of 170000 under nonreducing conditions and 135000 (α subunit) and 25000 (β subunit) under reducing conditions. GP IX had an apparent Mr of 22000 under both nonreducing and reducing conditions. Purified Gb Ib complex and GP Ib inhibited the ristocetin-mediated, human factor VIII/von Willebrand-factor-dependent and bovine factor VIII/von Willebrand-factor-dependent agglutination of washed human platelets suggesting the proteins had been isolated in functionally active form. GP Ibα had a similar amino acid composition to that previously reported for its proteolytic degradation product, glycocalicin. The amino acid compositions of GP Ibβ and GP IX were similar but showed marked differences in the levels of glutamic acid, alanine, histidine and arginine. The N-termini of GP Ibα and GP IX were blocked; GP Ibβ had the N-terminal sequence, Ile-Pro-Ala-Pro-. On crossed immunoelectrophoresis, both GP Ib and GP IX were found to occur in the same immunoprecipitin arc(s) whether the platelets had been solubilized in the absence or presence of the calcium-dependent protease inhibitor, leupeptin. Binding studies in platelet-rich plasma indicated a similar number of binding sites (x± SD) for three anti-(glycoprotein Ib complex) monoclonal antibodies: AN 51, epitope on GP Ibα (22000 ± 2700, n= 3), WM 23, epitope on GP Ibα (21000 ± 3400, n= 3), FMC 25, epitope on GP IX (20100 ± 2700, n= 3), and FMC 25 (Fab′)2 (27100 ± 800, n= 2). The combined evidence suggests that GP Ib is normally bound to GP IX and that the stoichiometry of this complex is 1:1.

https://febs.onlinelibrary.wiley.com/doi/epdf/10.1111/j.1432-1033.1985.tb09152.x

Purification and preliminary characterization of the glycoprotein Ib complex in the human platelet membrane

Drug disposition and drug hypersensitivity.

Additional glycoprotein defects in Bernard-Soulier's syndrome: confirmation of genetic basis by parental analysis

Drug-induced thrombocytopenia (DIT) is a relatively common clinical disorder. It is imperative to provide rapid identification and removal of the offending agent before clinically significant bleeding or, in the case of heparin, thrombosis occurs. DIT can be distinguished from idiopathic thrombocytopenic purpura, a bleeding disorder caused by thrombocytopenia not associated with a systemic disease, based on the history of drug ingestion or injection and laboratory findings. DIT disorders can be a consequence of decreased platelet production (bone marrow suppression) or accelerated platelet destruction (especially immune-mediated destruction).

Drug-induced thrombocytopenia.

Previous studies have shown that the receptor for quinine- and quinidine-dependent antibodies is not expressed on the surface of platelets from patients with the Bernard-Soulier (B-S) syndrome. We now report data to suggest that these platelets lack the receptor for these antibodies. Since B-S platelets are also missing 2 related components of the GP I complex, GP lb and glycocalicin, our findings suggest that the receptor for quinine- and quinidine-dependent antibodies may be associated with this complex on normal platelets. An antibody previously shown to be directed against a surface antigen that migrated in the GP I position on SDS-polyacrylamide gel electrophoresis specifically blocked the reaction of the receptor with quinine- or quinidine-dependent antibodies. Purified glycocalicin, however, lacked detectable receptor activity. In contrast, a mixture of GP lb and a putative structural analog of this glycoprotein (Mr 210,000) eluted from wheat germ affinity columns after chromatography of Triton X soluble preparations of platelets or membranes was shown to contain at least 80% of the total receptor activity. Our results strongly suggest that the receptor is associated with GP lb and/or its high m.w. structural analog(s).

J P Caen

Papers

Further studies of the human platelet receptor for quinine- and quinine-dependent antibodies.

patients with the gray platelet syndrome Specific protein and glycoprotein deficiencies in platelets isolated from two