Structural architecture of prothrombin in solution revealed by single molecule spectroscopy
Abstract
The coagulation factor prothrombin has a complex spatial organization of its modular assembly that comprises the N-terminal Gla domain, kringle-1, kringle-2, and the C-terminal protease domain connected by three intervening linkers. Here we use single molecule Förster resonance energy transfer to access the conformational landscape of prothrombin in solution and uncover structural features of functional significance that extend recent x-ray crystallographic analysis. Prothrombin exists in equilibrium between two alternative conformations, open and closed. The closed conformation predominates (70%) and features an unanticipated intramolecular collapse of Tyr93 in kringle-1 onto Trp547 in the protease domain that obliterates access to the active site and protects the zymogen from autoproteolytic conversion to thrombin. The open conformation (30%) is more susceptible to chymotrypsin digestion and autoactivation, and features a shape consistent with recent x-ray crystal structures. Small angle x-ray scattering measurements of prothrombin wild type stabilized 70% in the closed conformation and of the mutant Y93A stabilized 80% in the open conformation directly document two envelopes that differ 50 Å in length. These findings reveal important new details on the conformational plasticity of prothrombin in solution and the drastic structural difference between its alternative conformations. Prothrombin uses the intramolecular collapse of kringle-1 onto themore »
- Authors:
-
- Saint Louis Univ. School of Medicine, St. Louis, MO (United States)
- Saint Louis Univ. School of Medicine, St. Louis, MO (United States); Wroclaw Univ. of Technology, Wroclaw (Poland)
- 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), Basic Energy Sciences (BES)
- OSTI Identifier:
- 1369035
- Grant/Contract Number:
- AC02-06CH11357
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of Biological Chemistry
- Additional Journal Information:
- Journal Volume: 291; Journal Issue: 35; Journal ID: ISSN 0021-9258
- Publisher:
- American Society for Biochemistry and Molecular Biology
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; 59 BASIC BIOLOGICAL SCIENCES; enzyme kinetics; prothrombin; single-molecule biophysics; structure-function; thrombin
Citation Formats
Pozzi, Nicola, Bystranowska, Dominika, Zuo, Xiaobing, and Di Cera, Enrico. Structural architecture of prothrombin in solution revealed by single molecule spectroscopy. United States: N. p., 2016.
Web. doi:10.1074/jbc.M116.738310.
Pozzi, Nicola, Bystranowska, Dominika, Zuo, Xiaobing, & Di Cera, Enrico. Structural architecture of prothrombin in solution revealed by single molecule spectroscopy. United States. https://doi.org/10.1074/jbc.M116.738310
Pozzi, Nicola, Bystranowska, Dominika, Zuo, Xiaobing, and Di Cera, Enrico. Tue .
"Structural architecture of prothrombin in solution revealed by single molecule spectroscopy". United States. https://doi.org/10.1074/jbc.M116.738310. https://www.osti.gov/servlets/purl/1369035.
@article{osti_1369035,
title = {Structural architecture of prothrombin in solution revealed by single molecule spectroscopy},
author = {Pozzi, Nicola and Bystranowska, Dominika and Zuo, Xiaobing and Di Cera, Enrico},
abstractNote = {The coagulation factor prothrombin has a complex spatial organization of its modular assembly that comprises the N-terminal Gla domain, kringle-1, kringle-2, and the C-terminal protease domain connected by three intervening linkers. Here we use single molecule Förster resonance energy transfer to access the conformational landscape of prothrombin in solution and uncover structural features of functional significance that extend recent x-ray crystallographic analysis. Prothrombin exists in equilibrium between two alternative conformations, open and closed. The closed conformation predominates (70%) and features an unanticipated intramolecular collapse of Tyr93 in kringle-1 onto Trp547 in the protease domain that obliterates access to the active site and protects the zymogen from autoproteolytic conversion to thrombin. The open conformation (30%) is more susceptible to chymotrypsin digestion and autoactivation, and features a shape consistent with recent x-ray crystal structures. Small angle x-ray scattering measurements of prothrombin wild type stabilized 70% in the closed conformation and of the mutant Y93A stabilized 80% in the open conformation directly document two envelopes that differ 50 Å in length. These findings reveal important new details on the conformational plasticity of prothrombin in solution and the drastic structural difference between its alternative conformations. Prothrombin uses the intramolecular collapse of kringle-1 onto the active site in the closed form to prevent autoactivation. As a result, the open-closed equilibrium also defines a new structural framework for the mechanism of activation of prothrombin by prothrombinase.},
doi = {10.1074/jbc.M116.738310},
journal = {Journal of Biological Chemistry},
number = 35,
volume = 291,
place = {United States},
year = {Tue Jul 19 00:00:00 EDT 2016},
month = {Tue Jul 19 00:00:00 EDT 2016}
}
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