Improvement of the recombinant human coagulation factor IX expression by co-expression of a novel transcript of Drosophila γ carboxylase in a human cell line.

Biotechnology Letters
Solmaz Moniri Javadhesari, Alireza Zomorodipour

Abstract

Mammalian cells as the main host for production of human proteins are incapable of complete γ-carboxylation of over-expressed Vitamin K Dependent (VKD) proteins. The Drosophila γ-glutamyl carboxylase (DγC) has been shown to be more efficient than its human counterpart in γ-carboxylation of human substrates, in vitro. Considering the Drosophila γ-carboxylase (DγC) efficiency, in comparison with its human counterpart, for recognition and γ-carboxylation of a human substrate in vitro, we were determined to study the effect of the DγC on the hFIX expression in a mammalian cell line. With this aim, we examined co-expression of the DγC with the hFIX, in a human cell line. While the co-expression of a complete DγC cDNA reduced the hFIX expression, a truncated form of DγC could improve both the expression level (up to 1211 ng/106 cells/ml on the 4th day of post-transfection) and carboxylation of the expressed hFIX, significantly (p < 0.009). Our findings provided evidences for potential of a partial fragment of the DγC for improvement of the γ-carboxylation of a human substrate in a mammalian cell. Our experimental data, in accordance with in silico analysis suggested that the DγC C-terminal fragment, with the advantage of a Kozak-like...Continue Reading

References

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Datasets Mentioned

BETA
NT-037436.3

Methods Mentioned

BETA
PCR
transfection
enzyme linked immunosorbent assay
ELISA
transfections
antigen assay

Software Mentioned

SPSS
TMHMM
ORF
ATGpr
TMHMM Server
DyCint
Blast
CoSiDe
SignalP

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