Factor IX oligomerization underlies reduced activity upon disruption of physiological conditions

V. L. Simhadri, N. Hamasaki-Katagiri, S. C. Tseng, A. A. Bentley, R. Zichel, A. Y. Hershko, A. A. Komar, C. Kimchi-Sarfaty*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Coagulation factor IX (FIX) is a serine protease that plays a pivotal role in the blood coagulation cascade. FIX deficiency leads to a blood clotting disorder known as haemophilia B. FIX, synthesized as a prepro-peptide of 461 amino acids, is processed and secreted into plasma. The protein undergoes numerous modifications, including, but not limited to glycosylation, γ-carboxylation and disulphide bond formation. Upon processing and limited proteolysis, the protein is converted into an active protease. Under physiological conditions, the FIX zymogen is a monomer. The purpose of this work was to analyse the conditions that may affect FIX monomeric state and promote and/or reduce oligomerization. Using native gel electrophoresis and size exclusion chromatography, we found that under decreased pH and ionic strength conditions, the FIX zymogen can oligomerize, resulting in the formation of higher molecular weight species, with a concomitant reduction in specific activity. Similarly, FIX oligomers formed readily with low bovine serum albumin (BSA) concentrations; however, increased BSA concentrations impeded FIX oligomerization. We hypothesize that normal blood physiological conditions are critical for maintaining active FIX monomers. Under conditions of stress associated with acidosis, electrolyte imbalance and low albumin levels, FIX oligomerization is expected to take place thus leading to compromised activity. Furthermore, albumin, which is commonly used as a drug stabilizer, may enhance the efficacy of FIX biological drugs by reducing oligomerization.

Original languageEnglish
Pages (from-to)e157-e163
JournalHaemophilia
Volume20
Issue number2
DOIs
StatePublished - 2014
Externally publishedYes

Funding

FundersFunder number
American Heart Association13GRNT17070025

    Keywords

    • BeneFIX
    • Clotting factor
    • Factor IX
    • Mononine
    • Oligomer

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