The Experts below are selected from a list of 2016 Experts worldwide ranked by ideXlab platform
James H. Morrissey - One of the best experts on this subject based on the ideXlab platform.
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interactions between platelets and the coagulation system
2019Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Abstract Coagulation consists of a Cascade of enzymes (often with a regulatory cofactor), ultimately resulting in the cleavage of fibrinogen to fibrin, which polymerizes into a gel. Thrombin generated by the Clotting Cascade is also a potent platelet activator. Platelets contain many pro- and anticoagulant components of the blood coagulation Cascade. Some participants are secreted from platelet granules upon activation, some are transferred from the cytoplasm, and some are receptors on the platelet surface. Links between coagulation participants, polymerized fibrin and aggregated platelets aid in anchoring the growing clot to the vascular wall. Activated platelets provide a critical surface for assembly of the intrinsic tenase complex (which generates factor Xa) and the prothrombinase complex (which generates thrombin). This surface allows for binding of blood Clotting enzymes, cofactors, and substrates via calcium-dependent interactions between Clotting proteins and anionic phospholipids (particularly phosphatidylserine) in the platelet membrane.
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Lipid specificity of the membrane binding domain of coagulation factor X.
Journal of thrombosis and haemostasis : JTH, 2017Co-Authors: Melanie P. Muller, James H. Morrissey, Yan Wang, Emad TajkhorshidAbstract:Essentials Membrane-binding GLA domains of coagulation factors are essential for proper clot formation. Factor X (FX) is specific to phosphatidylserine (PS) lipids through unknown atomic-level interactions. Molecular dynamics simulations were used to develop the first membrane-bound model of FX-GLA. PS binding modes of FX-GLA were described, and potential PS-specific binding sites identified. SummaryBackground Factor X (FX) binds to cell membranes in a highly phospholipid-dependent manner and, in complex with tissue factor and factor VIIa (FVIIa), initiates the Clotting Cascade. Experimental information concerning the membrane-bound structure of FX with atomic resolution has remained elusive because of the fluid nature of cellular membranes. FX is known to bind preferentially to phosphatidylserine (PS). Objectives To develop the first membrane-bound model of the FX-GLA domain to PS at atomic level, and to identify PS-specific binding sites of the FX-GLA domain. Methods Molecular dynamics (MD) simulations were performed to develop an atomic-level model for the FX-GLA domain bound to PS bilayers. We utilized a membrane representation with enhanced lipid mobility, termed the highly mobile membrane mimetic (HMMM), permitting spontaneous membrane binding and insertion by FX-GLA in multiple 100-ns simulations. In 14 independent simulations, FX-GLA bound spontaneously to the membrane. The resulting membrane-bound models were converted from HMMM to conventional membrane and simulated for an additional 100 ns. Results The final membrane-bound FX-GLA model allowed for detailed characterization of the orientation, insertion depth and lipid interactions of the domain, providing insight into the molecular basis of its PS specificity. All binding simulations converged to the same configuration despite differing initial orientations. Conclusions Analysis of interactions between residues in FX-GLA and lipid-charged groups allowed for potential PS-specific binding sites to be identified. This new structural and dynamic information provides an additional step towards a full understanding of the role of atomic-level lipid–protein interactions in regulating the critical and complex Clotting Cascade.
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High-Resolution NMR Studies of Human Tissue Factor
2016Co-Authors: Kristin M. Nuzzio, James H. Morrissey, Eric D. Watt, John M. Boettcher, Joshua M. Gajsiewicz, Chad M. RienstraAbstract:In normal hemostasis, the blood Clotting Cascade is initiated when factor VIIa (fVIIa, other Clotting factors are named similarly) binds to the integral membrane protein, human tissue factor (TF). The TF/fVIIa complex in turn activates fX and fIX, eventually concluding with clot formation. Several X-ray crystal structures of the soluble extracellular domain of TF (sTF) exist; however, these structures are missing electron density in functionally relevant regions of the protein. In this context, NMR can provide complementary structural information as well as dynamic insights into enzyme activity. The resolution and sensitivity for NMR studies are greatly enhanced by the ability to prepare multiple milligrams of protein with various isotopic labeling patterns. Here, we demonstrate high-yield production of several isotopically labeled forms of recombinant sTF, allowing for high-resolution NMR studies both in the solid and solution state. We also report solution NMR spectra at sub-mM concentrations of sTF, ensuring the presence of dispersed monomer, as well as the first solid-state NMR spectra of sTF. Our improved sample preparation and precipitation conditions have enabled the acquisition of multidimensional NMR data sets for TF chemical shift assignment and provide a benchmark for TF structure elucidation.
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2013 scientific sessions sol sherry distinguished lecture in thrombosissignificance
Arteriosclerosis Thrombosis and Vascular Biology, 2015Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Polyphosphate is a highly anionic, linear polymer of inorganic phosphates that is found throughout biology, including in many infectious microorganisms. Recently, polyphosphate was discovered to be stored in a subset of the secretory granules of human platelets and mast cells, and to be secreted on activation of these cells. Work from our laboratory and others has now shown that polyphosphate is a novel, potent modulator of the blood Clotting and complement systems that likely plays roles in hemostasis, thrombosis, inflammation, and host responses to pathogens. Therapeutics targeting polyphosphate may have the potential to limit thrombosis with fewer hemorrhagic complications than conventional anticoagulant drugs that target essential proteases of the blood Clotting Cascade. # Significance {#article-title-93}
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2013 scientific sessions sol sherry distinguished lecture in thrombosis polyphosphate a novel modulator of hemostasis and thrombosis
Arteriosclerosis Thrombosis and Vascular Biology, 2015Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Polyphosphate is a highly anionic, linear polymer of inorganic phosphates that is found throughout biology, including in many infectious microorganisms. Recently, polyphosphate was discovered to be stored in a subset of the secretory granules of human platelets and mast cells, and to be secreted on activation of these cells. Work from our laboratory and others has now shown that polyphosphate is a novel, potent modulator of the blood Clotting and complement systems that likely plays roles in hemostasis, thrombosis, inflammation, and host responses to pathogens. Therapeutics targeting polyphosphate may have the potential to limit thrombosis with fewer hemorrhagic complications than conventional anticoagulant drugs that target essential proteases of the blood Clotting Cascade.
Stephanie A Smith - One of the best experts on this subject based on the ideXlab platform.
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interactions between platelets and the coagulation system
2019Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Abstract Coagulation consists of a Cascade of enzymes (often with a regulatory cofactor), ultimately resulting in the cleavage of fibrinogen to fibrin, which polymerizes into a gel. Thrombin generated by the Clotting Cascade is also a potent platelet activator. Platelets contain many pro- and anticoagulant components of the blood coagulation Cascade. Some participants are secreted from platelet granules upon activation, some are transferred from the cytoplasm, and some are receptors on the platelet surface. Links between coagulation participants, polymerized fibrin and aggregated platelets aid in anchoring the growing clot to the vascular wall. Activated platelets provide a critical surface for assembly of the intrinsic tenase complex (which generates factor Xa) and the prothrombinase complex (which generates thrombin). This surface allows for binding of blood Clotting enzymes, cofactors, and substrates via calcium-dependent interactions between Clotting proteins and anionic phospholipids (particularly phosphatidylserine) in the platelet membrane.
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2013 scientific sessions sol sherry distinguished lecture in thrombosissignificance
Arteriosclerosis Thrombosis and Vascular Biology, 2015Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Polyphosphate is a highly anionic, linear polymer of inorganic phosphates that is found throughout biology, including in many infectious microorganisms. Recently, polyphosphate was discovered to be stored in a subset of the secretory granules of human platelets and mast cells, and to be secreted on activation of these cells. Work from our laboratory and others has now shown that polyphosphate is a novel, potent modulator of the blood Clotting and complement systems that likely plays roles in hemostasis, thrombosis, inflammation, and host responses to pathogens. Therapeutics targeting polyphosphate may have the potential to limit thrombosis with fewer hemorrhagic complications than conventional anticoagulant drugs that target essential proteases of the blood Clotting Cascade. # Significance {#article-title-93}
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2013 scientific sessions sol sherry distinguished lecture in thrombosis polyphosphate a novel modulator of hemostasis and thrombosis
Arteriosclerosis Thrombosis and Vascular Biology, 2015Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Polyphosphate is a highly anionic, linear polymer of inorganic phosphates that is found throughout biology, including in many infectious microorganisms. Recently, polyphosphate was discovered to be stored in a subset of the secretory granules of human platelets and mast cells, and to be secreted on activation of these cells. Work from our laboratory and others has now shown that polyphosphate is a novel, potent modulator of the blood Clotting and complement systems that likely plays roles in hemostasis, thrombosis, inflammation, and host responses to pathogens. Therapeutics targeting polyphosphate may have the potential to limit thrombosis with fewer hemorrhagic complications than conventional anticoagulant drugs that target essential proteases of the blood Clotting Cascade.
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how it all starts initiation of the Clotting Cascade
Critical Reviews in Biochemistry and Molecular Biology, 2015Co-Authors: Stephanie A Smith, Richard J Travers, James H. MorrisseyAbstract:The plasma coagulation system in mammalian blood consists of a Cascade of enzyme activation events in which serine proteases activate the proteins (proenzymes and procofactors) in the next step of the Cascade via limited proteolysis. The ultimate outcome is the polymerization of fibrin and the activation of platelets, leading to a blood clot. This process is protective, as it prevents excessive blood loss following injury (normal hemostasis). Unfortunately, the blood Clotting system can also lead to unwanted blood clots inside blood vessels (pathologic thrombosis), which is a leading cause of disability and death in the developed world. There are two main mechanisms for triggering the blood Clotting, termed the tissue factor pathway and the contact pathway. Only one of these pathways (the tissue factor pathway) functions in normal hemostasis. Both pathways, however, are thought to contribute to thrombosis. An emerging concept is that the contact pathway functions in host pathogen defenses. This review focuses on how the initiation phase of the blood Clotting Cascade is regulated in both pathways, with a discussion of the contributions of these pathways to hemostasis versus thrombosis.
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2013 scientific sessions sol sherry distinguished lecture in thrombosis
Arteriosclerosis Thrombosis and Vascular Biology, 2015Co-Authors: Stephanie A Smith, James H. MorrisseyAbstract:Polyphosphate is a highly anionic, linear polymer of inorganic phosphates that is found throughout biology, including in many infectious microorganisms. Recently, polyphosphate was discovered to be stored in a subset of the secretory granules of human platelets and mast cells, and to be secreted on activation of these cells. Work from our laboratory and others has now shown that polyphosphate is a novel, potent modulator of the blood Clotting and complement systems that likely plays roles in hemostasis, thrombosis, inflammation, and host responses to pathogens. Therapeutics targeting polyphosphate may have the potential to limit thrombosis with fewer hemorrhagic complications than conventional anticoagulant drugs that target essential proteases of the blood Clotting Cascade.
Nigel Mackman - One of the best experts on this subject based on the ideXlab platform.
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Tissue Factor Past, Present, and Future
2015Co-Authors: Nigel Mackman, Mark TaubmanAbstract:This issue of Arteriosclerosis, Thrombosis, and Vascular Biology contains 4 reviews on tissue factor (TF) and 1 on tissue factor pathway inhibitor (TFPI). One review on TF will be published in a later issue. In this editorial, we will briefly revisit the major advances in the field, highlight some of the current controversies, and discuss some of the future challenges. TF (also known as tissue thromboplastin or coagulation factor III) was first identified as a constituent of tissue that when added to plasma activated the Clotting Cascade — hence the name tissue factor. TF was first purified in 1985,1 and this subsequently led to the cloning of the TF cDNA and gene. 2-5 In 1989 Drake and colleagues6 proposed that TF around blood vessels forms a “hemostatic envelope ” that initiates Clotting after vessel injury. The crystal structure of the extracellular domain of TF bound to Factor VIIa (FVIIa) was reported in 1996.7 In the same year it was discovered that inactivation of the mouse TF gene resulted in embryonic lethality.8-10 Taken together, these studies indicated that TF was essential for hemostasis. Activation of the Clotting Cascade leads to the generation of thrombin that cleaves fibrinogen to fibrin as well as activates platelets (Figure). In 1999, the late Yale Nemerson and colleagues11 reported that there was TF in blood of health
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Tissue Factor and Tissue Factor Pathway Inhibitor as Key Regulators of Global Hemostasis: Measurement of Their Levels
2015Co-Authors: In Coagulation Assays, Nigel Mackman, Raj S. Kasthuri, Sam L. Glover, Jeremiah Boles, Ph. DAbstract:The tissue factor (TF)/factor (F)VIIa complex is the primary initiator of coagulation in vivo. Tissue factor pathway inhibitor (TFPI) is the physiological inhibitor of the TF/FVIIa complex. Deficiencies of either TF or TFPI have not been reported in humans, and a complete absence of either of these two proteins in mice is embryonically lethal. To maintain normal hemostasis, levels of TF and TFPI need to be balanced. Increased levels of TF can overwhelm the inhibitory capacity of TFPI, resulting in thrombosis. Decreased levels of TF are associated with bleeding. Global assays of coagulation are defined as tests capable of evaluating all components of the Clotting Cascade that are present in plasma. In these tests the thrombogenic surface is either provided by platelets or exogenous phospholipids. Clotting assays currently used in clinical practice are not designed to measure endogenous levels of TF and TFPI. Therefore, there is a need to develop sensitive and specific assays for measuring levels of functional TF and TFPI in whole blood and plasma. These assays could be useful in patient management in many scenarios. Keywords Tissue factor; tissue factor pathway inhibitor; thrombosis; thromboelastograph
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microparticles in hemostasis and thrombosis
Circulation Research, 2011Co-Authors: Phillip A Owens, Nigel MackmanAbstract:Blood contains microparticles (MPs) derived from a variety of cell types, including platelets, monocytes, and endothelial cells. In addition, tumors release MPs into the circulation. MPs are formed from membrane blebs that are released from the cell surface by proteolytic cleavage of the cytoskeleton. All MPs are procoagulant because they provide a membrane surface for the assembly of components of the coagulation protease Cascade. Importantly, procoagulant activity is increased by the presence of anionic phospholipids, particularly phosphatidylserine (PS), and the procoagulant protein tissue factor (TF), which is the major cellular activator of the Clotting Cascade. High levels of platelet-derived PS + MPs are present in healthy individuals, whereas the number of TF + , PS + MPs is undetectable or very low. However, levels of PS + , TF + MPs are readily detected in a variety of diseases, and monocytes appear to be the primary cellular source. In cancer, PS + , TF + MPs are derived from tumors and may serve as a useful biomarker to identify patients at risk for venous thrombosis. This review will summarize our current knowledge of the role of procoagulant MPs in hemostasis and thrombosis.
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the role of tissue factor and factor viia in hemostasis
Anesthesia & Analgesia, 2009Co-Authors: Nigel MackmanAbstract:The coagulation system has been historically divided into three pathways: the extrinsic, intrinsic, and common pathways (Fig. 1).1–3 The tissue factor (TF): Factor VII/VIIa (FVII/FVIIa) complex was called the “extrinsic” pathway because an exogenous agent (i.e., TF) was required for activation of the Clotting factors in plasma. The TF:FVIIa complex is the key initiator of the coagulation protease Cascade and activates both FIX to FIXa and FX to FXa. This leads to the formation of low amounts of thrombin, which activates the cofactors FV and FVIII. The prothrombin time is determined by adding exogenous TF to plasma and is used to assess the activity of the extrinsic pathway. Components of the “intrinsic” pathway (FXII, FXI, FIX, and FVIII) are all present in blood.4 The tenase complex (FVIIIa:FIXa) plays a key role in amplifying the Clotting Cascade by activating FX to FXa. The activated partial prothrombin time uses kaolin or other negatively charged substances to activate FXII and is used to assess the activity of the intrinsic pathway. The prothrombinase complex (FVa:FXa) and thrombin are referred to as the common pathway. The prothrombinase complex activates prothrombin to thrombin, which is the central protease of the Clotting Cascade. Thrombin cleaves fibrinogen into soluble fibrin monomers that polymerize. Thrombin also activates the transglutaminase FXIII to FXIIIa that in turn cross-links soluble fibrin monomers into a fibrin matrix. Finally, thrombin activates platelets by cleavage of protease activated receptors.5 Figure 1 Simplified version of the Clotting Cascade. Tissue factor (TF) and FVIIa comprise the extrinsic pathway. FXIIa, FXIa, FIXa, and FVIIIa are members of the intrinsic pathway. The common pathway is made up of FVa, FXa, and thrombin. Thrombin cleaves fibrinogen, ... The coagulation Cascade is regulated by several anticoagulants. A kunitz-type serine protease inhibitor called TF pathway inhibitor inhibits the TF:FVIIa complex by forming a quaternary complex with FXa.6 Protein C is converted to activated protein C by a thrombin-thrombomodulin complex located on the surface of endothelial cells.7 Activated protein C in association with its cofactor protein S cleaves and inactivates the cofactors FV and FVIII. The primary target of the anticoagulant protein antithrombin is thrombin, although it can also inactivate other coagulation proteases in the Cascade, including FIXa, FXa, FXIa, and FXIIa.8
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the role of tissue factor and factor viia in hemostasis
Anesthesia & Analgesia, 2009Co-Authors: Nigel MackmanAbstract:Tissue factor (TF) is a transmembrane receptor for Factor VII/VIIa (FVII/VIIa). It is constitutively expressed by cells surrounding blood vessels. The endothelium physically separates this potent "activator" from its circulating ligand FVII/FVIIa and prevents inappropriate activation of the Clotting Cascade. Breakage of the endothelial barrier leads to exposure of extravascular TF and rapid activation of the Clotting Cascade. TF is also expressed in certain tissues, such as the heart and brain, and provides additional hemostatic protection to these tissues. Small amounts of TF are also present in blood in the form of microparticles, which are small membrane vesicles derived from activated and apoptotic cells. Levels of microparticle TF increase in a variety of diseases, such as sepsis and cancer, and this so-called "blood-borne" TF may contribute to thrombosis associated with these diseases. Recombinant FVIIa has been developed as an effective hemostatic drug for the treatment of hemophilia patients with inhibitory antibodies. In addition, it is used for patients with bleeding that do not respond to conventional therapy. However, the mechanism by which recombinant FVIIa restores hemostasis has not been clearly defined. In conclusion, the TF:FVIIa complex is essential for hemostasis and recombinant FVIIa is an effective hemostatic drug.
Sadaaki Iwanaga - One of the best experts on this subject based on the ideXlab platform.
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horseshoe crab acetyl group recognizing lectins involved in innate immunity are structurally related to fibrinogen
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Soutaro Gokudan, Sadaaki Iwanaga, Tatsushi Muta, Ryoko Tsuda, Kumiko Koori, Takeshi Kawahara, Noriaki Seki, Yoshimitsu Mizunoe, Sun Nyunt Wai, Shun-ichiro KawabataAbstract:We have characterized and cloned newly isolated lectins from hemolymph plasma of the horseshoe crab Tachypleus tridentatus, which we named tachylectins 5A and 5B (TLs-5). TLs-5 agglutinated all types of human erythrocytes and Gram-positive and Gram-negative bacteria. TLs-5 specifically recognize acetyl group-containing substances including noncarbohydrates; the acetyl group is required and is sufficient for recognition. TLs-5 enhanced the antimicrobial activity of a horseshoe crab-derived big defensin. cDNA sequences of TLs-5 indicated that they consist of a short N-terminal Cys-containing segment and a C-terminal fibrinogen-like domain with the highest sequence identity (51%) to that of mammalian ficolins. TLs-5, however, lack the collagenous domain found in a kind of “bouquet arrangement” of ficolins and collectins. Electron microscopy revealed that TLs-5 form two- to four-bladed propeller structures. The horseshoe crab is equipped with a unique functional homologue of vertebrate fibrinogen, coagulogen, as the target protein of the Clotting Cascade. Our observations clearly show that the horseshoe crab has fibrinogen-related molecules in hemolymph plasma and that they function as nonself-recognizing lectins. An ancestor of fibrinogen may have functioned as a nonself-recognizing protein.
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crystal structure of a coagulogen the Clotting protein from horseshoe crab a structural homologue of nerve growth factor
The EMBO Journal, 1996Co-Authors: Andreas Bergner, Sadaaki Iwanaga, Tatsushi Muta, Vaheh Oganessyan, Dieter Typke, Robert Huber, Wolfram BodeAbstract:Abstract The Clotting Cascade system of the horseshoe crab (Limulus) is involved in both haemostasis and host defence. The Cascade results in the conversion of coagulogen, a soluble protein, into an insoluble coagulin gel. The Clotting enzyme excises the fragment peptide C from coagulogen, giving rise to aggregation of the monomers. The crystal structure of coagulogen reveals an elongated molecule that embraces the helical peptide C fragment. Cleavage and removal of the peptide C would expose an extended hydrophobic cove, which could interact with the hydrophobic edge of a second molecule, leading to a polymeric fibre. The C-terminal half of the coagulogen molecule exhibits a striking topological similarity to the neurotrophin nerve growth factor (NGF), providing the first evidence for a neurotrophin fold in invertebrates. Similarities between coagulogen and Spatzle, the Drosophila ligand of the receptor Toll, suggest that the neurotrophin fold might be considered more ancient and widespread than previously realized.
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purified horseshoe crab factor g reconstitution and characterization of the 1 3 β d glucan sensitive serine protease Cascade
Journal of Biological Chemistry, 1995Co-Authors: Tatsushi Muta, Fuminori Tokunaga, Noriaki Seki, Ryuji Hashimoto, Yoshie Takaki, Toshio Oda, Atsufumi Iwanaga, Sadaaki IwanagaAbstract:Horseshoe crab hemocyte lysate responds to (1 → 3)-β-D-glucans, initiating an enzymatic Cascade, which culminates in clot formation.We have purified to homogeneity the serine protease zymogen factor G, which is directly activated by (1 → 3)-β-D-glucans and which initiates the hemolymph Clotting Cascade. Factor G is a heterodimeric protein composed of two noncovalently associated subunits α (72 kDa) and β (37 kDa). In the presence of (1 → 3)-β-D-glucans such as curdlan and paramylon, factor G is autocatalytically activated to an active serine protease named factor G. This activation is accompanied by limited proteolysis of both subunits: the 72-kDa subunit α is cleaved to 55-kDa and 17-kDa fragments, and the 37-kDa subunit β is shortened to 34 kDa. Longer incubations with (1 → 3)-β-D-glucans result in cleavage of the 55-kDa fragment to 46 kDa and the 34-kDa fragment to 32 kDa, with concomitant loss of amidase activity. Reconstitution experiments using purified proteins participating in the hemolymph Clotting Cascade demonstrate that factor G is capable of activating proClotting enzyme directly, resulting in the conversion of coagulogen to coagulin gel. Thus, purified factor G is shown to be the primary initiator of the (1 → 3)-β-D-glucan-sensitive coagulation pathway in the horseshoe crab hemocyte lysate.
Tatsushi Muta - One of the best experts on this subject based on the ideXlab platform.
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horseshoe crab acetyl group recognizing lectins involved in innate immunity are structurally related to fibrinogen
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Soutaro Gokudan, Sadaaki Iwanaga, Tatsushi Muta, Ryoko Tsuda, Kumiko Koori, Takeshi Kawahara, Noriaki Seki, Yoshimitsu Mizunoe, Sun Nyunt Wai, Shun-ichiro KawabataAbstract:We have characterized and cloned newly isolated lectins from hemolymph plasma of the horseshoe crab Tachypleus tridentatus, which we named tachylectins 5A and 5B (TLs-5). TLs-5 agglutinated all types of human erythrocytes and Gram-positive and Gram-negative bacteria. TLs-5 specifically recognize acetyl group-containing substances including noncarbohydrates; the acetyl group is required and is sufficient for recognition. TLs-5 enhanced the antimicrobial activity of a horseshoe crab-derived big defensin. cDNA sequences of TLs-5 indicated that they consist of a short N-terminal Cys-containing segment and a C-terminal fibrinogen-like domain with the highest sequence identity (51%) to that of mammalian ficolins. TLs-5, however, lack the collagenous domain found in a kind of “bouquet arrangement” of ficolins and collectins. Electron microscopy revealed that TLs-5 form two- to four-bladed propeller structures. The horseshoe crab is equipped with a unique functional homologue of vertebrate fibrinogen, coagulogen, as the target protein of the Clotting Cascade. Our observations clearly show that the horseshoe crab has fibrinogen-related molecules in hemolymph plasma and that they function as nonself-recognizing lectins. An ancestor of fibrinogen may have functioned as a nonself-recognizing protein.
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crystal structure of a coagulogen the Clotting protein from horseshoe crab a structural homologue of nerve growth factor
The EMBO Journal, 1996Co-Authors: Andreas Bergner, Sadaaki Iwanaga, Tatsushi Muta, Vaheh Oganessyan, Dieter Typke, Robert Huber, Wolfram BodeAbstract:Abstract The Clotting Cascade system of the horseshoe crab (Limulus) is involved in both haemostasis and host defence. The Cascade results in the conversion of coagulogen, a soluble protein, into an insoluble coagulin gel. The Clotting enzyme excises the fragment peptide C from coagulogen, giving rise to aggregation of the monomers. The crystal structure of coagulogen reveals an elongated molecule that embraces the helical peptide C fragment. Cleavage and removal of the peptide C would expose an extended hydrophobic cove, which could interact with the hydrophobic edge of a second molecule, leading to a polymeric fibre. The C-terminal half of the coagulogen molecule exhibits a striking topological similarity to the neurotrophin nerve growth factor (NGF), providing the first evidence for a neurotrophin fold in invertebrates. Similarities between coagulogen and Spatzle, the Drosophila ligand of the receptor Toll, suggest that the neurotrophin fold might be considered more ancient and widespread than previously realized.
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purified horseshoe crab factor g reconstitution and characterization of the 1 3 β d glucan sensitive serine protease Cascade
Journal of Biological Chemistry, 1995Co-Authors: Tatsushi Muta, Fuminori Tokunaga, Noriaki Seki, Ryuji Hashimoto, Yoshie Takaki, Toshio Oda, Atsufumi Iwanaga, Sadaaki IwanagaAbstract:Horseshoe crab hemocyte lysate responds to (1 → 3)-β-D-glucans, initiating an enzymatic Cascade, which culminates in clot formation.We have purified to homogeneity the serine protease zymogen factor G, which is directly activated by (1 → 3)-β-D-glucans and which initiates the hemolymph Clotting Cascade. Factor G is a heterodimeric protein composed of two noncovalently associated subunits α (72 kDa) and β (37 kDa). In the presence of (1 → 3)-β-D-glucans such as curdlan and paramylon, factor G is autocatalytically activated to an active serine protease named factor G. This activation is accompanied by limited proteolysis of both subunits: the 72-kDa subunit α is cleaved to 55-kDa and 17-kDa fragments, and the 37-kDa subunit β is shortened to 34 kDa. Longer incubations with (1 → 3)-β-D-glucans result in cleavage of the 55-kDa fragment to 46 kDa and the 34-kDa fragment to 32 kDa, with concomitant loss of amidase activity. Reconstitution experiments using purified proteins participating in the hemolymph Clotting Cascade demonstrate that factor G is capable of activating proClotting enzyme directly, resulting in the conversion of coagulogen to coagulin gel. Thus, purified factor G is shown to be the primary initiator of the (1 → 3)-β-D-glucan-sensitive coagulation pathway in the horseshoe crab hemocyte lysate.