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Title: Enhancing the anticoagulant profile of meizothrombin

Abstract

Meizothrombin is an active intermediate generated during the proteolytic activation of prothrombin to thrombin in the penultimate step of the coagulation cascade. Structurally, meizothrombin differs from thrombin because it retains the auxiliary Gla domain and two kringles. Functionally, meizothrombin shares with thrombin the ability to cleave procoagulant (fibrinogen), prothrombotic (PAR1) and anticoagulant (protein C) substrates, although its specificity toward fibrinogen and PAR1 is less pronounced. In this study we report information on the structural architecture of meizothrombin resolved by SAXS and single molecule FRET as an elongated arrangement of its individual domains. In addition, we show the properties of a meizothrombin construct analogous to the anticoagulant thrombin mutant W215A/E217A currently in Phase I for the treatment of thrombotic complications and stroke. The findings reveal new structural and functional aspects of meizothrombin that advance our understanding of a key intermediate of the prothrombin activation pathway.

Authors:
 [1];  [1];  [2];  [1];  [1]
  1. Saint Louis Univ. School of Medicine, St. Louis, MO (United States)
  2. Argonne National Lab. (ANL), Argonne, IL (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States)
Sponsoring Org.:
USDOE Office of Science (SC); American Heart Association; National Institutes of Health (NIH)
OSTI Identifier:
1504767
Grant/Contract Number:  
AC02-06CH11357; 15SDG25550094; HL049413; HL139554
Resource Type:
Accepted Manuscript
Journal Name:
Biomolecular Concepts
Additional Journal Information:
Journal Volume: 9; Journal Issue: 1; Journal ID: ISSN 1868-5021
Publisher:
de Gruyter
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; enzxyme specificity; protein engineering; thrombin

Citation Formats

Stojanovski, Bosko M., Pelc, Leslie A., Zuo, Xiaobing, Pozzi, Nicola, and Cera, Enrico Di. Enhancing the anticoagulant profile of meizothrombin. United States: N. p., 2018. Web. doi:10.1515/bmc-2018-0016.
Stojanovski, Bosko M., Pelc, Leslie A., Zuo, Xiaobing, Pozzi, Nicola, & Cera, Enrico Di. Enhancing the anticoagulant profile of meizothrombin. United States. doi:10.1515/bmc-2018-0016.
Stojanovski, Bosko M., Pelc, Leslie A., Zuo, Xiaobing, Pozzi, Nicola, and Cera, Enrico Di. Wed . "Enhancing the anticoagulant profile of meizothrombin". United States. doi:10.1515/bmc-2018-0016. https://www.osti.gov/servlets/purl/1504767.
@article{osti_1504767,
title = {Enhancing the anticoagulant profile of meizothrombin},
author = {Stojanovski, Bosko M. and Pelc, Leslie A. and Zuo, Xiaobing and Pozzi, Nicola and Cera, Enrico Di},
abstractNote = {Meizothrombin is an active intermediate generated during the proteolytic activation of prothrombin to thrombin in the penultimate step of the coagulation cascade. Structurally, meizothrombin differs from thrombin because it retains the auxiliary Gla domain and two kringles. Functionally, meizothrombin shares with thrombin the ability to cleave procoagulant (fibrinogen), prothrombotic (PAR1) and anticoagulant (protein C) substrates, although its specificity toward fibrinogen and PAR1 is less pronounced. In this study we report information on the structural architecture of meizothrombin resolved by SAXS and single molecule FRET as an elongated arrangement of its individual domains. In addition, we show the properties of a meizothrombin construct analogous to the anticoagulant thrombin mutant W215A/E217A currently in Phase I for the treatment of thrombotic complications and stroke. The findings reveal new structural and functional aspects of meizothrombin that advance our understanding of a key intermediate of the prothrombin activation pathway.},
doi = {10.1515/bmc-2018-0016},
journal = {Biomolecular Concepts},
number = 1,
volume = 9,
place = {United States},
year = {2018},
month = {12}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record

Figures / Tables:

Figure 1 Figure 1: Solution structures of prothrombin and Mz revealed by SAXS and smFRET measurements. (A) Reconstructed 3D envelopes of prothrombin (red) and Mz (blue) calculated from their respective scattering profiles. The maximal particle sizes (Dmax) for prothrombin and Mz calculated from the pair distance distribution functions (not shown) are 120more » Å and 160 Å, respectively. FRET efficiency histogram of Mz S525A labeled with the AF555/AF647 fluorophores at positions S120C/S478C measured in the absence (B) or presence(C) of saturating concentrations of H-D-Phe-Pro-Arg-p-nitroanilide (FPR). (D) Fractions of Mz molecules populating either the open (closed circles) or closed (open circles) conformations calculated at different concentrations of FPR. The Kd value for the interaction of Mz with FPR obtained from fitting the data into a single site model was 90±10 nM.« less

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Works referenced in this record:

The linker connecting the two kringles plays a key role in prothrombin activation
journal, May 2014

  • Pozzi, N.; Chen, Z.; Pelc, L. A.
  • Proceedings of the National Academy of Sciences, Vol. 111, Issue 21
  • DOI: 10.1073/pnas.1403779111

PAM: A Framework for Integrated Analysis of Imaging, Single-Molecule, and Ensemble Fluorescence Data
journal, April 2018


Interplay between conformational selection and zymogen activation
journal, March 2018

  • Chakraborty, Pradipta; Acquasaliente, Laura; Pelc, Leslie A.
  • Scientific Reports, Vol. 8, Issue 1
  • DOI: 10.1038/s41598-018-21728-9

How the Linker Connecting the Two Kringles Influences Activation and Conformational Plasticity of Prothrombin
journal, January 2016

  • Pozzi, Nicola; Chen, Zhiwei; Di Cera, Enrico
  • Journal of Biological Chemistry, Vol. 291, Issue 12
  • DOI: 10.1074/jbc.M115.700401

Identification of residues linked to the slow-->fast transition of thrombin.
journal, November 1995

  • Guinto, E. R.; Vindigni, A.; Ayala, Y. M.
  • Proceedings of the National Academy of Sciences, Vol. 92, Issue 24
  • DOI: 10.1073/pnas.92.24.11185

Rational Design of a Potent Anticoagulant Thrombin
journal, November 2000

  • Cantwell, Angelene M.; Di Cera, Enrico
  • Journal of Biological Chemistry, Vol. 275, Issue 51
  • DOI: 10.1074/jbc.C000751200

Protein C activation on endothelial cells by prothrombin activation products generated in situ: meizothrombin is a better protein C activator than α-thrombin
journal, October 1996

  • Hackeng, Tilman M.; Tans, Guido; Koppelman, Stefan J.
  • Biochemical Journal, Vol. 319, Issue 2
  • DOI: 10.1042/bj3190399

Cofactor Proteins in the Assembly and Expression of Blood Clotting Enzyme Complexes
journal, June 1988


Limiting prothrombin activation to meizothrombin is compatible with survival but significantly alters hemostasis in mice
journal, August 2016


Mechanism of the Anticoagulant Activity of Thrombin Mutant W215A/E217A
journal, July 2009

  • Gandhi, Prafull S.; Page, Michael J.; Chen, Zhiwei
  • Journal of Biological Chemistry, Vol. 284, Issue 36
  • DOI: 10.1074/jbc.M109.025403

The Thrombin Mutant W215A/E217A Shows Safe and Potent Anticoagulant and Antithrombotic Effects in Vivo
journal, June 2002

  • Gruber, Andras; Cantwell, Angelene M.; Di Cera, Enrico
  • Journal of Biological Chemistry, Vol. 277, Issue 31
  • DOI: 10.1074/jbc.C200237200

Autocatalytic Peptide Bond Cleavages in Prothrombin and Meizothrombin
journal, February 1998

  • Petrovan, Ramona J.; Govers-Riemslag, José W. P.; Nowak, Götz
  • Biochemistry, Vol. 37, Issue 5
  • DOI: 10.1021/bi971948h

The Anticoagulant Thrombin Mutant W215A/E217A Has a Collapsed Primary Specificity Pocket
journal, July 2004

  • Pineda, Agustin O.; Chen, Zhi-Wei; Caccia, Sonia
  • Journal of Biological Chemistry, Vol. 279, Issue 38
  • DOI: 10.1074/jbc.M407272200

WEDGE: an anticoagulant thrombin mutant produced by autoactivation
journal, November 2014

  • Wood, D. C.; Pelc, L. A.; Pozzi, N.
  • Journal of Thrombosis and Haemostasis, Vol. 13, Issue 1
  • DOI: 10.1111/jth.12774

The transition of prothrombin to thrombin
journal, June 2013

  • Krishnaswamy, S.
  • Journal of Thrombosis and Haemostasis, Vol. 11
  • DOI: 10.1111/jth.12217

Na+ binding to meizothrombin desF1
journal, October 2008

  • Papaconstantinou, M. E.; Gandhi, P. S.; Chen, Z.
  • Cellular and Molecular Life Sciences, Vol. 65, Issue 22
  • DOI: 10.1007/s00018-008-8502-7

Meizothrombin, a Major Product of Factor Xa-Catalyzed Prothrombin Activation
journal, December 1988


Factor XI Activation by Meizothrombin: Stimulation by Phospholipid Vesicles Containing both Phosphatidylserine and Phosphatidylethanolamine
journal, June 1997

  • Borne, Peter; Mosnier, Laurent; Tans, Guido
  • Thrombosis and Haemostasis, Vol. 78, Issue 01
  • DOI: 10.1055/s-0038-1657637

Structural Architecture of Prothrombin in Solution Revealed by Single Molecule Spectroscopy
journal, July 2016

  • Pozzi, Nicola; Bystranowska, Dominika; Zuo, Xiaobing
  • Journal of Biological Chemistry, Vol. 291, Issue 35
  • DOI: 10.1074/jbc.M116.738310

Analysis of the Kinetics of Prothrombin Activation and Evidence That Two Equilibrating Forms of Prothrombinase Are Involved in the Process
journal, December 2002

  • Brufatto, Nicole; Nesheim, Michael E.
  • Journal of Biological Chemistry, Vol. 278, Issue 9
  • DOI: 10.1074/jbc.M206413200

Structural basis of the activation and action of trypsin
journal, March 1978

  • Huber, Robert; Bode, Wolfram
  • Accounts of Chemical Research, Vol. 11, Issue 3
  • DOI: 10.1021/ar50123a006

Calcium Ion Modulation of Meizothrombin Autolysis at Arg-Asp and Catalytic Activity
journal, April 1996

  • Stevens, Willem K.; Côté, Hélène C. F.; MacGillivray, Ross T. A.
  • Journal of Biological Chemistry, Vol. 271, Issue 14
  • DOI: 10.1074/jbc.271.14.8062

Structure of prothrombin in the closed form reveals new details on the mechanism of activation
journal, February 2018


Thrombomodulin Structure and Function
journal, June 1997


Crystal Structure of Prothrombin Reveals Conformational Flexibility and Mechanism of Activation
journal, June 2013

  • Pozzi, Nicola; Chen, Zhiwei; Gohara, David W.
  • Journal of Biological Chemistry, Vol. 288, Issue 31
  • DOI: 10.1074/jbc.M113.466946

Relative antithrombotic and antihemostatic effects of protein C activator versus low-molecular-weight heparin in primates
journal, January 2007


Engineering Thrombin for Selective Specificity toward Protein C and PAR1
journal, April 2010

  • Marino, Francesca; Pelc, Leslie A.; Vogt, Austin
  • Journal of Biological Chemistry, Vol. 285, Issue 25
  • DOI: 10.1074/jbc.M110.119875

Determination of Domain Structure of Proteins from X-Ray Solution Scattering
journal, June 2001


Kinetic Dissection of the Pre-existing Conformational Equilibrium in the Trypsin Fold
journal, July 2015

  • Vogt, Austin D.; Chakraborty, Pradipta; Di Cera, Enrico
  • Journal of Biological Chemistry, Vol. 290, Issue 37
  • DOI: 10.1074/jbc.M115.675538

Mutations in Autolytic Loop-2 and at Asp 554 of Human Prothrombin That Enhance Protein C Activation by Meizothrombin
journal, February 2003

  • Koike, Hisashi; Okuda, Daiju; Morita, Takashi
  • Journal of Biological Chemistry, Vol. 278, Issue 17
  • DOI: 10.1074/jbc.M208220200

Binding of Substrate in Two Conformations to Human Prothrombinase Drives Consecutive Cleavage at Two Sites in Prothrombin
journal, October 2004

  • Orcutt, Steven J.; Krishnaswamy, Sriram
  • Journal of Biological Chemistry, Vol. 279, Issue 52
  • DOI: 10.1074/jbc.M410866200

    Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.