Browsing by Subject "fibrinogen"

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    The Effect of Free Amino Acids on Fibrin Formation
    (Office of the Vice Chancellor for Research, 2014-04-11) Dow, Anna M.; Johnson, Merrell A.; Rosen, Elliot D.; Ray, Bruce D.; Petrache, Horia I.
    Fibrinogen, a plasma protein, is a main component of blood clot formation. In the event of an injury, blood loss is hindered through a process that forms a thrombus by conversion of fibrinogen to fibrin induced by activation of the enzyme thrombin. The fibrin network structure depends on the concentration of thrombin, as expected, but is also highly influenced by its environment during formation. In particular, we find that fibrin formation is altered in the presence of zwitterions. Zwitterions are dipolar molecules, typically highly polarizable, exhibiting both a positive and a negative charge depending on the pH of the solution. Amino acids are highly abundant zwitterions in biological materials. In this study we measured by visible/UV-spectroscopy the effects of various types of amino acids on the rate of fibrin network formation. We show that the electrical charge and type of amino acid, such as lysine, glycine and arginine, inhibits or promotes formation of fibrin networks. Such an ability to decrease or increase the rate of coagulation can be valuable in the treatment of patients suffering from hemostatic and thrombotic disorders.
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    Small Angle Scattering Of Large Protein Units Under Osmotic Stress
    (2020-05) Palacio, Luis A.; Petrache, Horia I.; Cheng, Ruihua; Joglekar, Yogesh N.; Liu, Jing; Wassall, Stephen R.
    Large protein molecules are abundant in biological cells but are very difficult to study in physiological conditions due to molecular disorder. For large proteins, most structural information is obtained in crystalline states which can be achieved in certain conditions at very low temperature. X-ray and neutron crystallography methods can then be used for determination of crystalline structures at atomic level. However, in solution at room or physiological temperatures such highly resolved descriptions cannot be obtained except in very few cases. Scattering methods that can be used to study this type of structures at room temperature include small-angle x-ray and neutron scattering. These methods are used here to study two distinct proteins that are both classified as glycoproteins, which are a large class of proteins with diverse biological functions. In this study, two specific plasma glycoproteins were used: Fibrinogen (340 kDa) and Alpha 1-Antitrypsin or A1AT (52 kDa). These proteins have been chosen based on the fact that they have a propensity to form very large molecular aggregates due to their tendency to polymerize. One goal of this project is to show that for such complex structures, a combination of scattering methods that include SAXS, SANS, and DLS can address important structural and interaction questions despite the fact that atomic resolution cannot be obtained as in crystallography. A1AT protein has been shown to have protective roles of lung cells against emphysema, while fibrinogen is a major factor in the blood clotting process. A systematic approach to study these proteins interactions with lipid membranes and other proteins, using contrast-matching small-angle neutron scattering (SANS), small angle x-ray scattering (SAXS) and dynamic light scattering (DLS), is presented here. A series of structural reference points for each protein in solution were determined by performing measurements under osmotic stress controlled by the addition of polyethylene glycol-1,500 MW (PEG 1500) in the samples. Osmotic pressure changes the free energy of the molecular mixture and has consequences on the structure and the interaction of molecular aggregates. In particular, the measured radius of gyration (Rg) for A1AT shows a sharp structural transition when the concentration of PEG 1500 is between 33 wt% and 36 wt%. Similarly, a significant structural change was observed for fibrinogen when the concentration of PEG 1500 was above 40 wt%. This analysis is applied to a study of A1AT interacting with lipid membranes and to a study of fibrinogen polymerization in the presence of the enzyme thrombin, which catalyzes the formation of blood clots. The experimental approach presented here and the applications to specific questions show that an appropriate combination of scattering methods can produce useful information on the behavior and the interactions of large protein systems in physiological conditions despite the lower resolution compared to crystallography.
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    Strong Binding of Platelet Integrin αIIbβ3 to Fibrin Clots: Potential Target to Destabilize Thrombi
    (Nature Publishing group, 2017-10-11) Höök, Peter; Litvinov, Rustem I.; Kim, Oleg V.; Xu, Shixin; Xu, Zhiliang; Bennett, Joel S.; Alber, Mark S.; Weisel, John W.; Medicine, School of Medicine
    The formation of platelet thrombi is determined by the integrin αIIbβ3-mediated interactions of platelets with fibrinogen and fibrin. Blood clotting in vivo is catalyzed by thrombin, which simultaneously induces fibrinogen binding to αIIbβ3 and converts fibrinogen to fibrin. Thus, after a short time, thrombus formation is governed by αIIbβ3 binding to fibrin fibers. Surprisingly, there is little understanding of αIIbβ3 interaction with fibrin polymers. Here we used an optical trap-based system to measure the binding of single αIIbβ3 molecules to polymeric fibrin and compare it to αIIbβ3 binding to monomeric fibrin and fibrinogen. Like αIIbβ3 binding to fibrinogen and monomeric fibrin, we found that αIIbβ3 binding to polymeric fibrin can be segregated into two binding regimes, one with weaker rupture forces of 30–60 pN and a second with stronger rupture forces >60 pN that peaked at 70–80 pN. However, we found that the mechanical stability of the bimolecular αIIbβ3-ligand complexes had the following order: fibrin polymer > fibrin monomer > fibrinogen. These quantitative differences reflect the distinct specificity and underlying molecular mechanisms of αIIbβ3-mediated reactions, implying that targeting platelet interactions with fibrin could increase the therapeutic indices of antithrombotic agents by focusing on the destabilization of thrombi rather than the prevention of platelet aggregation.