Advertisement

Purification and Autoactivation Method for Recombinant Coagulation Factor VII

  • Vladimir GranovskiEmail author
  • Marcela C. C. Freitas
  • Mario Soares Abreu-Neto
  • Dimas T. Covas
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1674)

Abstract

Recombinant coagulation factor VII is a very important and complex protein employed for treatment of hemophiliac patients (hemophilia A/B) who develop inhibitors antibodies to conventional treatments (FVIII and FIX). The rFVII is a glycosylated molecule and circulates in plasma as zymogen of 50 kDa. When activated the molecule is cleaved to 20–30 kDa and has a half-life of about 3 h, needing to be processed fast and efficiently until freeze-drying. Here, we describe a very simple and fast purification sequence for rFVII using affinity FVII Select resin and a dialysis system that can be easily scaled up.

Key words

Recombinant coagulation factor VII Affinity chromatography FVII autoactivation rFVII purification methodology 

References

  1. 1.
    Franchini M, Mannucci PM (2012) Past, present and future of hemophilia: a narrative review. Orphanet J Rare Dis 7:24CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Hedner U (2015) Recombinant activated factor VII: 30 years of research and innovation. Blood Rev Suppl 1:S4–S8CrossRefGoogle Scholar
  3. 3.
    Yoshihiro M, Takamiya O (2012) The expression of recombinant factor VII variants is influenced by different cells for transfection using expression studies. Rinsho Byori 60(3):193–200PubMedGoogle Scholar
  4. 4.
    Eigenbrot C (2002) Structure, function, and activation of coagulation factor VII. Curr Protein Pept Sci 3(3):287–299CrossRefPubMedGoogle Scholar
  5. 5.
    Bharadwaj D, Iino M, Kontoyianni M, Smith KJ, Foster DC, Kisiel W (1996) Factor VII central. A novel mutation in the catalytic domain that reduces tissue factor binding, impairs activation by factor Xa, and abolishes amidolytic and coagulant activity. J Biol Chem 271(48):30685–30691CrossRefPubMedGoogle Scholar
  6. 6.
    Tiede A (2015) Half-life extended factor VIII for the treatment of hemophilia A. J Thromb Haemost Suppl 1:S176–S179CrossRefGoogle Scholar
  7. 7.
    Pedersen AH, Lund-Hansen T, Bisgaard-Frantzen H, Olsen F, Petersen LC (1989) Autoactivation of human recombinant coagulation factor VII. Biochemistry 28(24):9331–9336CrossRefPubMedGoogle Scholar
  8. 8.
    Covas DT, Freitas MCC, Picanço-Castro V, Swiech K (2015) Processo de Produção de Fator VII de coagulação sanguínea e fator VII de coagulação sanguínea. Instituto Nacional da Propriedade Industrial (INPI) BR1020150123345Google Scholar

Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  • Vladimir Granovski
    • 1
    Email author
  • Marcela C. C. Freitas
    • 1
    • 2
  • Mario Soares Abreu-Neto
    • 2
  • Dimas T. Covas
    • 1
    • 2
  1. 1.Center for Cell-based Therapy CTCRibeirão Preto Medical School, University of São PauloRibeirão PretoBrazil
  2. 2.Regional Blood Center of Ribeirão PretoRibeirão Preto Medical School, University of São PauloRibeirão PretoBrazil

Personalised recommendations