lucia morales | Universidad Americana (Nicaragua) (original) (raw)
Address: Managua, Managua, Nicaragua
less
Related Authors
Università Cattolica del Sacro Cuore (Catholic University of the Sacred Heart)
Uploads
Papers by lucia morales
Journal of Thrombosis and Haemostasis, 2007
Summary. Background: von Willebrand factor (VWF) does not interact with circulating platelets unl... more Summary. Background: von Willebrand factor (VWF) does not interact with circulating platelets unless it is induced to expose the binding site for platelet glycoprotein (GP)Ibα in the A1 domain by high shear stress, immobilization, and/or a modulator. Previous studies have implied indirectly that the A2 domain may be involved in regulating A1–GPIbα binding. Objective and methods: Because the relationship between the A1 and A2 domains has not been defined, we have investigated the effect of the A2 domain on the binding activity of the A1 domain using recombinant A domain polypeptides, multimeric VWF, and monoclonal antibodies (mAb). Results: The A2 domain polypeptide bound specifically to the immobilized A1 domain polypeptide or full-length VWF, with half-maximal binding being obtained at 60 or 168 nm, respectively. This A1–A2 interaction was inhibited by mAb against the A2 or A1 domain and by the A1 domain polypeptide. The A2 domain polypeptide effectively blocked GPIbα-mediated platelet adhesion under high flow conditions. The A2 domain polypeptide specifically recognizes the GPIbα-binding conformation in the A1 domain, as it only interacted with VWF activated by the modulator ristocetin or immobilized VWF. Furthermore, in contrast to plasma VWF, the ultra-large (UL)VWF multimers or a recombinant VWF–A1A2A3 polypeptide containing a gain-of-function mutation (R1308 L) of type 2B von Willebrand disease bound to the A2 domain polypeptide without the need for ristocetin. Conclusions: The recombinant A2 domain polypeptide specifically binds to the active conformation of the A1 domain in VWF and effectively blocks the interaction with platelet GPIbα under high-flow conditions.
Journal of Thrombosis and Haemostasis, 2006
Summary. Background: It is established that the A3 domain in von Willebrand factor (VWF) contains... more Summary. Background: It is established that the A3 domain in von Willebrand factor (VWF) contains the major collagen-binding site. However, there are conflicting reports describing the capacity of the A1 domain to interact with collagen types I and III. Methods: In this study, we have used recombinant VWF-A1 polypeptides, as well as conformation-specific monoclonal antibodies (mAb), to analyze the A1–collagen interaction. Results: The A1 domain bound to collagen with Kd ∼ 8.0 nm and this binding was blocked by the mAb 6G1, which blocks the interaction between ristocetin and VWF. In addition, collagen-bound A1 protein was able to support flow-dependent adhesion of platelets, demonstrating that the binding sites for collagen and glycoprotein (GP)Ib are different. Analysis with two conformation-specific mAb demonstrated that the structure of the A1 domain changed as a result of the binding to collagen. In contrast, the antibodies failed to detect conformational change in the G1324S mutant (type 2M von Willebrand disease). Thus, direct binding to collagen induces a change in the structural conformation within the VWF-A1 domain, and the G1324S substitution prevents this conformational change. Conclusion: This study has shown that the isolated A1 domain can simultaneously bind to collagen and platelet GPIb, supporting platelet adhesion under high-flow conditions. In addition, this study has used mAb to demonstrate that the binding of the isolated A1 domain or full-length VWF to collagen is accompanied by a conformational change in A1 domain.
Journal of Thrombosis and Haemostasis, 2007
Summary. Background: von Willebrand factor (VWF) does not interact with circulating platelets unl... more Summary. Background: von Willebrand factor (VWF) does not interact with circulating platelets unless it is induced to expose the binding site for platelet glycoprotein (GP)Ibα in the A1 domain by high shear stress, immobilization, and/or a modulator. Previous studies have implied indirectly that the A2 domain may be involved in regulating A1–GPIbα binding. Objective and methods: Because the relationship between the A1 and A2 domains has not been defined, we have investigated the effect of the A2 domain on the binding activity of the A1 domain using recombinant A domain polypeptides, multimeric VWF, and monoclonal antibodies (mAb). Results: The A2 domain polypeptide bound specifically to the immobilized A1 domain polypeptide or full-length VWF, with half-maximal binding being obtained at 60 or 168 nm, respectively. This A1–A2 interaction was inhibited by mAb against the A2 or A1 domain and by the A1 domain polypeptide. The A2 domain polypeptide effectively blocked GPIbα-mediated platelet adhesion under high flow conditions. The A2 domain polypeptide specifically recognizes the GPIbα-binding conformation in the A1 domain, as it only interacted with VWF activated by the modulator ristocetin or immobilized VWF. Furthermore, in contrast to plasma VWF, the ultra-large (UL)VWF multimers or a recombinant VWF–A1A2A3 polypeptide containing a gain-of-function mutation (R1308 L) of type 2B von Willebrand disease bound to the A2 domain polypeptide without the need for ristocetin. Conclusions: The recombinant A2 domain polypeptide specifically binds to the active conformation of the A1 domain in VWF and effectively blocks the interaction with platelet GPIbα under high-flow conditions.
Journal of Thrombosis and Haemostasis, 2006
Summary. Background: It is established that the A3 domain in von Willebrand factor (VWF) contains... more Summary. Background: It is established that the A3 domain in von Willebrand factor (VWF) contains the major collagen-binding site. However, there are conflicting reports describing the capacity of the A1 domain to interact with collagen types I and III. Methods: In this study, we have used recombinant VWF-A1 polypeptides, as well as conformation-specific monoclonal antibodies (mAb), to analyze the A1–collagen interaction. Results: The A1 domain bound to collagen with Kd ∼ 8.0 nm and this binding was blocked by the mAb 6G1, which blocks the interaction between ristocetin and VWF. In addition, collagen-bound A1 protein was able to support flow-dependent adhesion of platelets, demonstrating that the binding sites for collagen and glycoprotein (GP)Ib are different. Analysis with two conformation-specific mAb demonstrated that the structure of the A1 domain changed as a result of the binding to collagen. In contrast, the antibodies failed to detect conformational change in the G1324S mutant (type 2M von Willebrand disease). Thus, direct binding to collagen induces a change in the structural conformation within the VWF-A1 domain, and the G1324S substitution prevents this conformational change. Conclusion: This study has shown that the isolated A1 domain can simultaneously bind to collagen and platelet GPIb, supporting platelet adhesion under high-flow conditions. In addition, this study has used mAb to demonstrate that the binding of the isolated A1 domain or full-length VWF to collagen is accompanied by a conformational change in A1 domain.