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Effects of prothrombin on the individual activated protein C-mediated cleavages of coagulation factor Va.

Tran, Sinh LU ; Norström, Eva LU and Dahlbäck, Björn LU (2008) In Journal of Biological Chemistry 283(11). p.6648-6655
Abstract
The factor Va (FVa) inactivation by activated protein C (APC), mediated by cleavages at Arg306 and Arg506 in FVa, is inhibited by both factor Xa (FXa) and prothrombin. While FXa is known to specifically inhibit the Arg506 cleavage, the effect of prothrombin has not been confined to one cleavage site. We used recombinant FV variants, FV:506Q/679Q and FV:306Q/679Q, to investigate the effect of prothrombin on the individual cleavage sites. The APC-mediated FVa inhibition was monitored by a prothrombinase-based FVa-assay and apparent first order rate constants were calculated for each of the cleavage sites both in the presence and absence of prothrombin. Prothrombin impaired cleavages at both Arg306 and Arg506 and the inhibition correlated... (More)
The factor Va (FVa) inactivation by activated protein C (APC), mediated by cleavages at Arg306 and Arg506 in FVa, is inhibited by both factor Xa (FXa) and prothrombin. While FXa is known to specifically inhibit the Arg506 cleavage, the effect of prothrombin has not been confined to one cleavage site. We used recombinant FV variants, FV:506Q/679Q and FV:306Q/679Q, to investigate the effect of prothrombin on the individual cleavage sites. The APC-mediated FVa inhibition was monitored by a prothrombinase-based FVa-assay and apparent first order rate constants were calculated for each of the cleavage sites both in the presence and absence of prothrombin. Prothrombin impaired cleavages at both Arg306 and Arg506 and the inhibition correlated with a delayed appearance of proteolytic products on Western blots. Almost complete inhibition was obtained at around 3 microM prothrombin, whereas half-maximal inhibition was obtained at 0.7 microM prothrombin. After cleavage of prothrombin by thrombin, the inhibitory activity was lost. The inhibitory effect of prothrombin on APC-mediated inhibition of FVa was seen both in the presence and absence of protein S but in particular for the Arg306 sites, it was more pronounced in the presence of protein S. Thus, prothrombin inhibition of APC inactivation of FVa appears to be due to both impaired APC function and decreased APC-cofactor function of protein S. In conclusion, FVa being part of the prothrombinase complex is protected from APC by both FXa and prothrombin. Release of products of prothrombin activation from the prothrombinase complex would alleviate the protection allowing APC-mediated inactivation of FVa. (Less)
Please use this url to cite or link to this publication:
author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Biological Chemistry
volume
283
issue
11
pages
6648 - 6655
publisher
ASBMB
external identifiers
  • pmid:18198180
  • wos:000253779600006
  • scopus:43749109436
ISSN
1083-351X
DOI
10.1074/jbc.M708036200
language
English
LU publication?
yes
id
593de843-0b4b-4b4d-a73b-6a8832009e20 (old id 1021338)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/18198180?dopt=Abstract
date added to LUP
2008-02-13 15:16:55
date last changed
2017-07-23 04:52:38
@article{593de843-0b4b-4b4d-a73b-6a8832009e20,
  abstract     = {The factor Va (FVa) inactivation by activated protein C (APC), mediated by cleavages at Arg306 and Arg506 in FVa, is inhibited by both factor Xa (FXa) and prothrombin. While FXa is known to specifically inhibit the Arg506 cleavage, the effect of prothrombin has not been confined to one cleavage site. We used recombinant FV variants, FV:506Q/679Q and FV:306Q/679Q, to investigate the effect of prothrombin on the individual cleavage sites. The APC-mediated FVa inhibition was monitored by a prothrombinase-based FVa-assay and apparent first order rate constants were calculated for each of the cleavage sites both in the presence and absence of prothrombin. Prothrombin impaired cleavages at both Arg306 and Arg506 and the inhibition correlated with a delayed appearance of proteolytic products on Western blots. Almost complete inhibition was obtained at around 3 microM prothrombin, whereas half-maximal inhibition was obtained at 0.7 microM prothrombin. After cleavage of prothrombin by thrombin, the inhibitory activity was lost. The inhibitory effect of prothrombin on APC-mediated inhibition of FVa was seen both in the presence and absence of protein S but in particular for the Arg306 sites, it was more pronounced in the presence of protein S. Thus, prothrombin inhibition of APC inactivation of FVa appears to be due to both impaired APC function and decreased APC-cofactor function of protein S. In conclusion, FVa being part of the prothrombinase complex is protected from APC by both FXa and prothrombin. Release of products of prothrombin activation from the prothrombinase complex would alleviate the protection allowing APC-mediated inactivation of FVa.},
  author       = {Tran, Sinh and Norström, Eva and Dahlbäck, Björn},
  issn         = {1083-351X},
  language     = {eng},
  number       = {11},
  pages        = {6648--6655},
  publisher    = {ASBMB},
  series       = {Journal of Biological Chemistry},
  title        = {Effects of prothrombin on the individual activated protein C-mediated cleavages of coagulation factor Va.},
  url          = {http://dx.doi.org/10.1074/jbc.M708036200},
  volume       = {283},
  year         = {2008},
}