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Zaverio M. Ruggeri - One of the best experts on this subject based on the ideXlab platform.
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genetic deletion of platelet Glycoprotein Ib Alpha but not its extracellular domain protects from atherosclerosis
Thrombosis and Haemostasis, 2014Co-Authors: Ekaterina K Koltsova, Zaverio M. Ruggeri, Prithu Sundd, Alessandro Zarpellon, Hui Ouyang, Zbigniew Mikulski, Antonella Zampolli, Klaus LeyAbstract:The pathogenesis of atherosclerosis involves the interplay of haematopoietic, stromal and endothelial cells. Platelet interactions with endothelium and leukocytes are pivotal for atherosclerosis promotion. Glycoprotein (GP) Ibα is the ligand-binding subunit of the platelet GPIb-IX-V receptor complex; its deficiency causes the Bernard-Soulier syndrome (BSS), characterised by absent platelet GPIb-IX-V, macrothrombocytopenia and bleeding. We designed this study to determine the role of platelet GPIbα in the pathogenesis of atherosclerosis using two unique knockout models. Ldlr-/- mice were reconstituted with wild-type (wt), GPIbα-/- (lacks GPIbα) or chimeric IL-4R/GPIbα-Tg (lacks GPIbα extracellular domain) bone marrow and assayed for atherosclerosis development after feeding with pro-atherogenic “western diet”. Here, we report that Ldlr-/- mice reconstituted with GPIbα-/- bone marrow developed less atherosclerosis compared to wt controls; accompanied by augmented accumulation of pro-inflammatory CD11b+ and CD11c+ myeloid cells, reduced oxLDL uptake and decreased TNF and IL 12p35 gene expression in the aortas. Flow cytometry and live cell imaging in whole blood-perfused microfluidic chambers revealed reduced platelet-monocyte aggregates in GPIbα-/- mice, which resulted in decreased monocyte activation. Interestingly, Ldlr -/- mice reconstituted with IL-4R/GPIbα-Tg bone marrow, producing less abnormal platelets, showed atherosclerotic lesions similar to wt mice. Platelet interaction with blood monocytes and accumulation of myeloid cells in the aortas were also essentially unaltered. Moreover, only complete GPIbα ablation altered platelet microparticles and CCL5 chemokine production. Thus, atherosclerosis reduction in mice lacking GPIbα may not result from the defective GPIbα-ligand binding, but more likely is a consequence of functional defects of GPIbα-/- platelets and reduced blood platelet counts.
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specific synergy of multiple substrate receptor interactions in platelet thrombus formation under flow
Cell, 1998Co-Authors: Brian Savage, Fanny Almusjacobs, Zaverio M. RuggeriAbstract:We have used confocal videomicroscopy in real time to delineate the adhesive interactions supporting platelet thrombus formation on biologically relevant surfaces. Type I collagen fIbrils exposed to flowing blood adsorb von Willebrand factor (vWF), to which platelets become initially tethered with continuous surface translocation mediated by the membrane Glycoprotein Ib Alpha. This step is essential at high wall shear rates to allow subsequent irreversIble adhesion and thrombus growth mediated by the integrins Alpha2beta1 and Alpha(IIb)beta3. On subendothelial matrix, endogenous vWF and adsorbed plasma vWF synergistically initiate platelet recruitment, and Alpha2beta1 remains key along with Alpha(IIb)beta3 for normal thrombus development at all but low shear rates. Thus, hemodynamic forces and substrate characteristics define the platelet adhesion pathways leading to thrombogenesis.
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localization and characterization of an Alpha thrombin binding site on platelet Glycoprotein Ib Alpha
Journal of Biological Chemistry, 1994Co-Authors: L De Marco, M Mazzucato, A Masotti, Zaverio M. RuggeriAbstract:Glycoprotein (GP) Ib Alpha is required for expression of the highest affinity Alpha-thrombin-binding site on platelets, possIbly contrIbuting to platelet activation through a pathway involving cleavage of a specific receptor. This function may be important for the initiation of hemostasis and may also play a role in the development of pathological vascular occlusion. We have now identified a discrete sequence in the extracytoplasmic domain of GP Ib Alpha, including residues 271-284 of the mature protein, which appears to be part of the high affinity Alpha-thrombin-binding site. Synthetic peptidyl mimetics of this sequence inhIbit Alpha-thrombin binding to GP Ib as well as platelet activation and aggregation induced by subnanomolar concentrations of the agonist; they also inhIbit Alpha-thrombin binding to purified glycocalicin, the isolated extracytoplasmic portion of GP Ib Alpha. The inhIbitory peptides interfere with the clotting of fIbrinogen by Alpha-thrombin but not with the amidolytic activity of the enzyme on a small synthetic substrate, a finding compatIble with the concept that the identified GP Ib Alpha sequence interacts with the anion-binding exosite of Alpha-thrombin but not with its active proteolytic site. The crucial structural elements of this sequence necessary for thrombin binding appear to be a cluster of negatively charged residues as well as three tyrosine residues that, in the native protein, may be sulfated. GP Ib Alpha has no significant overall sequence homology with the thrombin inhIbitor, hirudin, nor with the specific thrombin receptor on platelets; all three molecules, however, possess a distinct region rich in negatively charged residues that appear to be involved in thrombin binding. This may represent a case of convergent evolution of unrelated proteins for high affinity interaction with the same ligand.
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expression of the phenotypic abnormality of platelet type von willebrand disease in a recombinant Glycoprotein Ib Alpha fragment
Journal of Clinical Investigation, 1993Co-Authors: M Murata, Zaverio M. Ruggeri, Susan Russell, Jerry WareAbstract:The platelet GP Ib-IX receptor supports platelet adhesion and activation by binding to vWf in the exposed subendothelial matrix. An abnormal GP Ib-IX complex exists in platelet-type or pseudo-von Willebrand disease and has a characteristic increased affinity for soluble vWf resulting in impaired hemostatic function due to the removal of larger vWf multimers from the circulation. Genetic studies within an afflicted family have demonstrated that the disease is linked to a Gly233-->Val amino acid substitution within the Alpha-subunit of the oligomeric GP Ib-IX complex (Miller, J.L., D. Cunningham, V.A. Lyle, and C. L. Finch. 1991. Proc. Natl. Acad. Sci. USA. 88:4761-4765). To evaluate the functional consequences of this mutation, we constructed a recombinant analogue of the Alpha-subunit of GP Ib containing Val233. Experiments comparing molecules with either Gly233 or Val233 revealed that the Val substitution generates a molecule with increased affinity for vWf. The recombinant fragment reproduces the functional abnormality of the GP Ib-IX complex in platelet-type von Willebrand disease, thus establishing the molecular basis of the bleeding disorder within this family. Moreover, it becomes apparent that structural elements responsIble for the regulation of hemostasis through modulation of vWf affinity for platelets reside within the Alpha-subunit of the GP Ib-IX complex.
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expression of human platelet Glycoprotein Ib Alpha in transgenic mice
Journal of Biological Chemistry, 1993Co-Authors: Jerry Ware, Susan Russell, Patrizia Marchese, Zaverio M. RuggeriAbstract:Platelets are cytoplasmic fragments of megakaryocytes and, therefore, their membrane proteins cannot be manipulated by expression methods in culture. To overcome this limitation, we have expressed human Glycoprotein (GP) Ib Alpha in transgenic mouse megakaryocytes and found that it was present on the surface of platelets associated with the mouse GP Ib beta subunit. This finding demonstrates that assembly of the heterooligomeric GP Ib complex occurs through mechanisms conserved across species. In contrast, the receptor function of GP Ib exhIbited restricted species specificity, since only the chimeric complex containing human GP Ib Alpha bound human von Willebrand factor and supported platelet aggregation mediated by ristocetin. These studies demonstrate the transgenic engineering of a platelet adhesion receptor under control of the human GP Ib Alpha promoter and illustrate a new approach to manipulate platelet receptors and study structure-function relationships in hemostasis and thrombosis.
Robert I. Handin - One of the best experts on this subject based on the ideXlab platform.
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Transient expression of recombinant Glycoprotein Ib Alpha polypeptides in COS cells that inhIbit von Willebrand factor binding to the platelet Glycoprotein Ib/IX complex.
Thrombosis and haemostasis, 1992Co-Authors: Eefke J Petersen, Robert I. HandinAbstract:The cDNA encoding the Glycoprotein Ib Alpha polypeptide has been expressed in COS cells. Transfection with full-length cDNA and a cDNA truncated at an internal XbaI site produced a recombinant polypeptide doublet with estimated molecular weights of 48 and 46 kDa (rGpIb Alpha L318) which could be resolved into a single band of molecular weight 36 kDa following digestion with endoglycosidase F. A portion of the truncated polypeptide was retained in the endoplasmic reticulum of COS cells and slowly released. A second fraction was rapidly secreted into COS cell-conditioned medium and could be used for functional studies. Soluble rGpIb Alpha L318 harvested from COS cell-conditioned medium inhIbited ristocetin-dependent binding of [125I]-vWF to fixed washed human platelets (IC50 20 nM). Binding was inhIbited by reduction and alkylation of "rGp1b Alpha L318" suggesting the need for a critical disulfide bond to maintain biological activity of the recombinant polypeptide. We conclude that the recombinant polypeptides produced by transfection of GpIb Alpha cDNA into heterologous cells are secreted, soluble, and can inhIbit vWF binding to platelet GpIb/IX.
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Functional analysis of a recombinant Glycoprotein Ib Alpha polypeptide which inhIbits von Willebrand factor binding to the platelet Glycoprotein Ib-IX complex and to collagen.
The Journal of biological chemistry, 1992Co-Authors: Miguel A. Cruz, Eefke J Petersen, S M Turci, Robert I. HandinAbstract:Abstract By deletion mutagenesis and transient expression in COS cells, a 96-amino acid hydrophilic sequence in the Glycoprotein Ib Alpha polypeptide located between L220 and L318 was identified which appeared to contain its von Willebrand factor- (vWF) binding site. The cDNA encoding this fragment was then expressed in Escherichia coli and purified from the bacterial cell lysate. The recombinant polypeptide, rGpIb Alpha Q221-L318, was monomeric and had an apparent molecular weight of 14,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It inhIbited both ristocetin-induced binding of 125I-vWF to fixed washed platelets and ristocetin-induced platelet agglutination. The recombinant polypeptide also inhIbited the binding of 125I-vWF to immobilized type I and III collagen. InhIbition of 125I-vWF binding to platelets and collagen was dose-dependent, with IC50 values of 500 and 200 nM rGpIb Alpha Q221-L318, respectively. Fifty % inhIbition of ristocetin-induced platelet agglutination required 500 nM rGpIb Alpha Q221-L318. Although rGpIb Alpha Q221-L318 inhIbited vWF binding to collagen it did not, itself, bind to collagen-coated surfaces. Reduction of the disulfide bond between C248 and C264 abolished activity. 125I-rGpIb Alpha Q221-L318 bound directly to GpIb/IX sites on multimeric vWF. These studies document that a portion of the sequence between Q221 and L318 is needed for recognition and binding to vWF and that binding requires an intact disulfide bond between C248 and C264. The binding of this recombinant polypeptide to vWF multimers inhIbits vWF interaction with two important substrates, platelet GpIb/IX and collagen.
M Murata - One of the best experts on this subject based on the ideXlab platform.
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platelet Glycoprotein Ib Alpha polymorphisms affect the interaction with von willebrand factor under flow conditions
British Journal of Haematology, 2005Co-Authors: Yumiko Matsubara, M Murata, Tomohiro Hayashi, Keijiro Suzuki, Yosuke Okamura, Makoto Handa, Hiroaki Ishihara, Toshiro Shibano, Yasuo IkedaAbstract:Interaction of platelet Glycoprotein (GP) IbAlpha with von Willebrand factor (VWF) is essential for thrombus formation, particularly under high shear conditions. Previous case-control studies indicated that two GPIb Alpha polymorphisms, (145)Thr/Met and/or variable number (1-4) tandem repeats of 13 amino-acid sequences, are associated with arterial thrombosis. The (145)Met-allele and the 3R- or 4R-allele is associated with increased risk. However, there is little clear experimental data to support this association. To elucidate the functional effects of these polymorphisms, we prepared recombinant GPIb Alpha fragments and tested them in vitro. The dissociation constants of ristocetin-induced (125)I-labelled VWF binding to two forms of soluble recombinant GPIb Alpha [(1)His-(302)Ala, either (145)Thr (145T) or (145)Met (145M)] were not different. Four types of Chinese hamster ovary cells expressing full-length GPIb Alpha beta/IX, 145T with one repeat (T1R), 145M with one repeat (M1R), 145T with four repeats (T4R), and 145M with four repeats (M4R), were prepared, and cell interactions with immobilized-VWF were examined under various shear conditions. The cell rolling velocity of M4R under a shear condition of 114/s was significantly slower than that of T1R. Intermediate values were obtained with M1R and T4R. The results suggest that M4R interacts more strongly with VWF under flow conditions.
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expression of the phenotypic abnormality of platelet type von willebrand disease in a recombinant Glycoprotein Ib Alpha fragment
Journal of Clinical Investigation, 1993Co-Authors: M Murata, Zaverio M. Ruggeri, Susan Russell, Jerry WareAbstract:The platelet GP Ib-IX receptor supports platelet adhesion and activation by binding to vWf in the exposed subendothelial matrix. An abnormal GP Ib-IX complex exists in platelet-type or pseudo-von Willebrand disease and has a characteristic increased affinity for soluble vWf resulting in impaired hemostatic function due to the removal of larger vWf multimers from the circulation. Genetic studies within an afflicted family have demonstrated that the disease is linked to a Gly233-->Val amino acid substitution within the Alpha-subunit of the oligomeric GP Ib-IX complex (Miller, J.L., D. Cunningham, V.A. Lyle, and C. L. Finch. 1991. Proc. Natl. Acad. Sci. USA. 88:4761-4765). To evaluate the functional consequences of this mutation, we constructed a recombinant analogue of the Alpha-subunit of GP Ib containing Val233. Experiments comparing molecules with either Gly233 or Val233 revealed that the Val substitution generates a molecule with increased affinity for vWf. The recombinant fragment reproduces the functional abnormality of the GP Ib-IX complex in platelet-type von Willebrand disease, thus establishing the molecular basis of the bleeding disorder within this family. Moreover, it becomes apparent that structural elements responsIble for the regulation of hemostasis through modulation of vWf affinity for platelets reside within the Alpha-subunit of the GP Ib-IX complex.
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genetic and structural characterization of an amino acid dimorphism in Glycoprotein Ib Alpha involved in platelet transfusion refractoriness
Blood, 1992Co-Authors: M Murata, Jerry Ware, Fumihiro Ishida, Kenichi Furihata, Susan Russell, Zaverio M. RuggeriAbstract:The platelet-specific alloantigen, SIba, located within the Alpha- subunit of the Glycoprotein (GP) Ib-IX membrane receptor, has been found to be involved in the pathogenesis of platelet transfusion refractoriness. We have identified the existence of a naturally occurring threonine/methionine dimorphism at position 145 of the GPIb Alpha sequence, and determined that the SIba antigen corresponds to the molecule containing methionine145. The diallelic codons can be detected by restriction enzyme analysis of amplified genomic DNA fragments from the GPIb Alpha gene. Evaluation of 61 healthy blood donors showed that the allele frequencies are 89% and 11% for the threonine145 and methionine145 codons, respectively. A positive correlation exists between platelet reactivity with the anti-SIba antIbody and the presence of a methionine145-encoding allele. Moreover, recombinant expression of two soluble GPIb Alpha fragments differing only at residue 145, provided definitive evidence that the human anti-SIba antIbody reacts only with the molecule containing methionine145. These results explain the structural basis of the SIba human alloantigen system and define screening methodologies useful in transfusion medicine to match donor and recipient platelets accordingly.
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Site-directed mutagenesis of a soluble recombinant fragment of platelet Glycoprotein Ib Alpha demonstrating negatively charged residues involved in von Willebrand factor binding.
The Journal of biological chemistry, 1991Co-Authors: M Murata, Jerry Ware, Zaverio M. RuggeriAbstract:We have expressed in mammalian cells a fragment (residues 1-302) of the Alpha chain of platelet Glycoprotein (GP) Ib containing the von Willebrand factor- (vWF) binding site. The secreted soluble protein had an apparent molecular mass of 45 kDa and reacted with conformation-dependent monoclonal antIbodies that bind only to native GP Ib, thus demonstrating its proper folding. After insolubilization on nitrocellulose membrane, the recombinant GP Ib Alpha fragment bound soluble vWF in the presence of ristocetin or botrocetin with a dissociation constant similar to that exhIbited by GP Ib.IX complex on platelets. Moreover, the interaction was blocked by anti-GP Ib monoclonal antIbodies known to inhIbit vWF binding to platelets. The sequence of GP Ib Alpha between residues 269-287 has a strong net negative charge due to the presence of 10 glutamic or aspartic acid residues; 5 of these are contained in the sequence of a synthetic peptide (residues 251-279) previously shown to inhIbit vWF-platelet interaction. In order to evaluate the possIble functional role of these acidic residues, we employed site-directed mutagenesis to express two mutant GP Ib Alpha fragments containing asparagine or glutamine instead of aspartic or glutamic acid, respectively. Mutant 1, with substitutions between residues 251-279, failed to bind vWF whether in the presence of ristocetin or botrocetin; in contrast, vWF binding to Mutant 2, with substitutions between residues 280-302, was nearly normal in the presence of ristocetin, but markedly decreased in the presence of botrocetin. Thus, mammalian cells transfected with a truncated cDNA sequence encoding the amino-terminal domain of GP Ib Alpha synthesize a fully functional vWF-binding site; acidic residues in the sequence 252-287 are essential for normal function.
Jerry Ware - One of the best experts on this subject based on the ideXlab platform.
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controlling elements of platelet Glycoprotein Ib Alpha expression
Comptes Rendus De L Academie Des Sciences Serie Iii-sciences De La Vie-life Sciences, 1996Co-Authors: Jerry Ware, Y Hashimoto, B Zieger, Susan R RussellAbstract:Our objectives are to define and characterize molecular events of megakaryocytopoiesis and platelet production by analyzing the in vivo expression of platelet Glycoprotein (GP) receptors. Our studies target the platelet GP Ib-IX-V complex since the congenital absence of the receptor results in the Bernard-Soulier syndrome, a condition linking the expression of the complex to normal platelet morphogenesis. We have previously descrIbed the generation of a transgenic mouse colony expressing the Alpha-subunit (GP Ib Alpha) of the human platelet GP Ib-IX-V complex. These studies established methodologies to manipulate GP Ib-IX-V on the platelet surface and examine unique aspects of hemostasis, megakaryocytopoiesis, platelet structure and platelet release. Our recent studies have defined the genetic elements supporting the megakaryocytic-expression of GP Ib Alpha. These results are defining essential cis-acting elements responsIble for the expression of GP Ib Alpha and are providing insights into molecular events coinciding with the release of normal platelets into the bloodstream.
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expression of the phenotypic abnormality of platelet type von willebrand disease in a recombinant Glycoprotein Ib Alpha fragment
Journal of Clinical Investigation, 1993Co-Authors: M Murata, Zaverio M. Ruggeri, Susan Russell, Jerry WareAbstract:The platelet GP Ib-IX receptor supports platelet adhesion and activation by binding to vWf in the exposed subendothelial matrix. An abnormal GP Ib-IX complex exists in platelet-type or pseudo-von Willebrand disease and has a characteristic increased affinity for soluble vWf resulting in impaired hemostatic function due to the removal of larger vWf multimers from the circulation. Genetic studies within an afflicted family have demonstrated that the disease is linked to a Gly233-->Val amino acid substitution within the Alpha-subunit of the oligomeric GP Ib-IX complex (Miller, J.L., D. Cunningham, V.A. Lyle, and C. L. Finch. 1991. Proc. Natl. Acad. Sci. USA. 88:4761-4765). To evaluate the functional consequences of this mutation, we constructed a recombinant analogue of the Alpha-subunit of GP Ib containing Val233. Experiments comparing molecules with either Gly233 or Val233 revealed that the Val substitution generates a molecule with increased affinity for vWf. The recombinant fragment reproduces the functional abnormality of the GP Ib-IX complex in platelet-type von Willebrand disease, thus establishing the molecular basis of the bleeding disorder within this family. Moreover, it becomes apparent that structural elements responsIble for the regulation of hemostasis through modulation of vWf affinity for platelets reside within the Alpha-subunit of the GP Ib-IX complex.
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expression of human platelet Glycoprotein Ib Alpha in transgenic mice
Journal of Biological Chemistry, 1993Co-Authors: Jerry Ware, Susan Russell, Patrizia Marchese, Zaverio M. RuggeriAbstract:Platelets are cytoplasmic fragments of megakaryocytes and, therefore, their membrane proteins cannot be manipulated by expression methods in culture. To overcome this limitation, we have expressed human Glycoprotein (GP) Ib Alpha in transgenic mouse megakaryocytes and found that it was present on the surface of platelets associated with the mouse GP Ib beta subunit. This finding demonstrates that assembly of the heterooligomeric GP Ib complex occurs through mechanisms conserved across species. In contrast, the receptor function of GP Ib exhIbited restricted species specificity, since only the chimeric complex containing human GP Ib Alpha bound human von Willebrand factor and supported platelet aggregation mediated by ristocetin. These studies demonstrate the transgenic engineering of a platelet adhesion receptor under control of the human GP Ib Alpha promoter and illustrate a new approach to manipulate platelet receptors and study structure-function relationships in hemostasis and thrombosis.
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genetic and structural characterization of an amino acid dimorphism in Glycoprotein Ib Alpha involved in platelet transfusion refractoriness
Blood, 1992Co-Authors: M Murata, Jerry Ware, Fumihiro Ishida, Kenichi Furihata, Susan Russell, Zaverio M. RuggeriAbstract:The platelet-specific alloantigen, SIba, located within the Alpha- subunit of the Glycoprotein (GP) Ib-IX membrane receptor, has been found to be involved in the pathogenesis of platelet transfusion refractoriness. We have identified the existence of a naturally occurring threonine/methionine dimorphism at position 145 of the GPIb Alpha sequence, and determined that the SIba antigen corresponds to the molecule containing methionine145. The diallelic codons can be detected by restriction enzyme analysis of amplified genomic DNA fragments from the GPIb Alpha gene. Evaluation of 61 healthy blood donors showed that the allele frequencies are 89% and 11% for the threonine145 and methionine145 codons, respectively. A positive correlation exists between platelet reactivity with the anti-SIba antIbody and the presence of a methionine145-encoding allele. Moreover, recombinant expression of two soluble GPIb Alpha fragments differing only at residue 145, provided definitive evidence that the human anti-SIba antIbody reacts only with the molecule containing methionine145. These results explain the structural basis of the SIba human alloantigen system and define screening methodologies useful in transfusion medicine to match donor and recipient platelets accordingly.
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Site-directed mutagenesis of a soluble recombinant fragment of platelet Glycoprotein Ib Alpha demonstrating negatively charged residues involved in von Willebrand factor binding.
The Journal of biological chemistry, 1991Co-Authors: M Murata, Jerry Ware, Zaverio M. RuggeriAbstract:We have expressed in mammalian cells a fragment (residues 1-302) of the Alpha chain of platelet Glycoprotein (GP) Ib containing the von Willebrand factor- (vWF) binding site. The secreted soluble protein had an apparent molecular mass of 45 kDa and reacted with conformation-dependent monoclonal antIbodies that bind only to native GP Ib, thus demonstrating its proper folding. After insolubilization on nitrocellulose membrane, the recombinant GP Ib Alpha fragment bound soluble vWF in the presence of ristocetin or botrocetin with a dissociation constant similar to that exhIbited by GP Ib.IX complex on platelets. Moreover, the interaction was blocked by anti-GP Ib monoclonal antIbodies known to inhIbit vWF binding to platelets. The sequence of GP Ib Alpha between residues 269-287 has a strong net negative charge due to the presence of 10 glutamic or aspartic acid residues; 5 of these are contained in the sequence of a synthetic peptide (residues 251-279) previously shown to inhIbit vWF-platelet interaction. In order to evaluate the possIble functional role of these acidic residues, we employed site-directed mutagenesis to express two mutant GP Ib Alpha fragments containing asparagine or glutamine instead of aspartic or glutamic acid, respectively. Mutant 1, with substitutions between residues 251-279, failed to bind vWF whether in the presence of ristocetin or botrocetin; in contrast, vWF binding to Mutant 2, with substitutions between residues 280-302, was nearly normal in the presence of ristocetin, but markedly decreased in the presence of botrocetin. Thus, mammalian cells transfected with a truncated cDNA sequence encoding the amino-terminal domain of GP Ib Alpha synthesize a fully functional vWF-binding site; acidic residues in the sequence 252-287 are essential for normal function.
Schulten Klaus - One of the best experts on this subject based on the ideXlab platform.
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Flow-Induced beta-Hairpin Folding of the Glycoprotein Ib Alpha beta-Switch
biophysical journal, 2010Co-Authors: Zou Xueqing, Liu Yanxin, Chen Zhongzhou, Ines Cardenas-jiron Gloria, Schulten KlausAbstract:Flow-induced shear has been identified as a regulatory driving force in blood clotting. Shear induces beta-hairpin folding of the Glycoprotein Ib Alpha beta-switch which increases affinity for binding to the von Willebrand factor, a key step in blood clot formation and wound healing. Through 2.1-mu s molecular dynamics simulations, we investigate the kinetics of flow-induced beta-hairpin folding. Simulations sampling different flow velocities reveal that under flow, beta-hairpin folding is initiated by hydrophobic collapse, followed by interstrand hydrogen-bond formation and turn formation. Adaptive biasing force simulations are employed to determine the free energy required for extending the unfolded beta-switch from a loop to an elongated state. Lattice and freely jointed chain models illustrate how the folding rate depends on the entropic and enthalpic energy, the latter controlled by flow. The results reveal that the free energy landscape of the beta-switch has two stable conformations imprinted on it, namely, loop and hairpin with flow inducing a transition between the two.BiophysicsSCI(E)PubMed7ARTICLE41182-11919