MASP-1 Induced Clotting - The First Model of Prothrombin Activation by MASP-1

Lorenz Jenny, J. Dobó, P. Gál, Verena Schroeder

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Mannan-binding lectin-associated serine protease-1 (MASP-1), a protein of the complement lectin pathway, resembles thrombin in terms of structural features and substrate specificity. Due to its interplay with several coagulation factors, it has the ability to induce fibrin clot formation independent of the usual coagulation activation pathways. We have recently shown that MASP-1 activates prothrombin and identified arginine (R) 155, R271, and R393 as potential cleavage sites. FXa cleaves R320 instead of R393, and thrombin cleaves R155 and R284 in prothrombin. Here we have used three arginine-to-glutamine mutants of prothrombin, R271Q, R320Q, R393Q and the serine-to-alanine active site mutant S525A to investigate in detail the mechanism of MASP-1 mediated prothrombin activation. Prothrombin wildtype and mutants were digested with MASP-1 and the cleavage products were analysed by SDS-PAGE and N-terminal sequencing. A functional clotting assay was performed by thrombelastography. We have found that MASP-1 activates prothrombin via two simultaneous pathways, either cleaving at R271 or R393 first. Both pathways result in the formation of several active alternative thrombin species. Functional studies confirmed that both R393 and R320 are required for prothrombin activation by MASP-1, whereas R155 is not considered to be an important cleavage site in this process. In conclusion, we have described for the first time a detailed model of prothrombin activation by MASP-1.

Original languageEnglish
Article numbere0144633
JournalPLoS One
Volume10
Issue number12
DOIs
Publication statusPublished - Dec 1 2015

Fingerprint

Mannose-Binding Protein-Associated Serine Proteases
Mannose-Binding Lectin
prothrombin
Serine Proteases
Prothrombin
serine proteinases
lectins
Chemical activation
thrombin
Thrombin
coagulation
mutants
arginine
Arginine
Mannose-Binding Lectin Complement Pathway
Thrombelastography
Blood Coagulation Factors
fibrin
substrate specificity
Substrate Specificity

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

MASP-1 Induced Clotting - The First Model of Prothrombin Activation by MASP-1. / Jenny, Lorenz; Dobó, J.; Gál, P.; Schroeder, Verena.

In: PLoS One, Vol. 10, No. 12, e0144633, 01.12.2015.

Research output: Contribution to journalArticle

@article{dcda9d277dd14c16b00dea60fd02cfee,
title = "MASP-1 Induced Clotting - The First Model of Prothrombin Activation by MASP-1",
abstract = "Mannan-binding lectin-associated serine protease-1 (MASP-1), a protein of the complement lectin pathway, resembles thrombin in terms of structural features and substrate specificity. Due to its interplay with several coagulation factors, it has the ability to induce fibrin clot formation independent of the usual coagulation activation pathways. We have recently shown that MASP-1 activates prothrombin and identified arginine (R) 155, R271, and R393 as potential cleavage sites. FXa cleaves R320 instead of R393, and thrombin cleaves R155 and R284 in prothrombin. Here we have used three arginine-to-glutamine mutants of prothrombin, R271Q, R320Q, R393Q and the serine-to-alanine active site mutant S525A to investigate in detail the mechanism of MASP-1 mediated prothrombin activation. Prothrombin wildtype and mutants were digested with MASP-1 and the cleavage products were analysed by SDS-PAGE and N-terminal sequencing. A functional clotting assay was performed by thrombelastography. We have found that MASP-1 activates prothrombin via two simultaneous pathways, either cleaving at R271 or R393 first. Both pathways result in the formation of several active alternative thrombin species. Functional studies confirmed that both R393 and R320 are required for prothrombin activation by MASP-1, whereas R155 is not considered to be an important cleavage site in this process. In conclusion, we have described for the first time a detailed model of prothrombin activation by MASP-1.",
author = "Lorenz Jenny and J. Dob{\'o} and P. G{\'a}l and Verena Schroeder",
year = "2015",
month = "12",
day = "1",
doi = "10.1371/journal.pone.0144633",
language = "English",
volume = "10",
journal = "PLoS One",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "12",

}

TY - JOUR

T1 - MASP-1 Induced Clotting - The First Model of Prothrombin Activation by MASP-1

AU - Jenny, Lorenz

AU - Dobó, J.

AU - Gál, P.

AU - Schroeder, Verena

PY - 2015/12/1

Y1 - 2015/12/1

N2 - Mannan-binding lectin-associated serine protease-1 (MASP-1), a protein of the complement lectin pathway, resembles thrombin in terms of structural features and substrate specificity. Due to its interplay with several coagulation factors, it has the ability to induce fibrin clot formation independent of the usual coagulation activation pathways. We have recently shown that MASP-1 activates prothrombin and identified arginine (R) 155, R271, and R393 as potential cleavage sites. FXa cleaves R320 instead of R393, and thrombin cleaves R155 and R284 in prothrombin. Here we have used three arginine-to-glutamine mutants of prothrombin, R271Q, R320Q, R393Q and the serine-to-alanine active site mutant S525A to investigate in detail the mechanism of MASP-1 mediated prothrombin activation. Prothrombin wildtype and mutants were digested with MASP-1 and the cleavage products were analysed by SDS-PAGE and N-terminal sequencing. A functional clotting assay was performed by thrombelastography. We have found that MASP-1 activates prothrombin via two simultaneous pathways, either cleaving at R271 or R393 first. Both pathways result in the formation of several active alternative thrombin species. Functional studies confirmed that both R393 and R320 are required for prothrombin activation by MASP-1, whereas R155 is not considered to be an important cleavage site in this process. In conclusion, we have described for the first time a detailed model of prothrombin activation by MASP-1.

AB - Mannan-binding lectin-associated serine protease-1 (MASP-1), a protein of the complement lectin pathway, resembles thrombin in terms of structural features and substrate specificity. Due to its interplay with several coagulation factors, it has the ability to induce fibrin clot formation independent of the usual coagulation activation pathways. We have recently shown that MASP-1 activates prothrombin and identified arginine (R) 155, R271, and R393 as potential cleavage sites. FXa cleaves R320 instead of R393, and thrombin cleaves R155 and R284 in prothrombin. Here we have used three arginine-to-glutamine mutants of prothrombin, R271Q, R320Q, R393Q and the serine-to-alanine active site mutant S525A to investigate in detail the mechanism of MASP-1 mediated prothrombin activation. Prothrombin wildtype and mutants were digested with MASP-1 and the cleavage products were analysed by SDS-PAGE and N-terminal sequencing. A functional clotting assay was performed by thrombelastography. We have found that MASP-1 activates prothrombin via two simultaneous pathways, either cleaving at R271 or R393 first. Both pathways result in the formation of several active alternative thrombin species. Functional studies confirmed that both R393 and R320 are required for prothrombin activation by MASP-1, whereas R155 is not considered to be an important cleavage site in this process. In conclusion, we have described for the first time a detailed model of prothrombin activation by MASP-1.

UR - http://www.scopus.com/inward/record.url?scp=84955612357&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84955612357&partnerID=8YFLogxK

U2 - 10.1371/journal.pone.0144633

DO - 10.1371/journal.pone.0144633

M3 - Article

VL - 10

JO - PLoS One

JF - PLoS One

SN - 1932-6203

IS - 12

M1 - e0144633

ER -