Decline in baculovirus-expressed recombinant protein production with increasing cell density is strongly correlated to impairment of virus replication and mRNA expression

Huynh, Hoai T., Tran, Trinh T. B., Chan, Leslie C. L., Nielsen, Lars K. and Reid, Steven (2013) Decline in baculovirus-expressed recombinant protein production with increasing cell density is strongly correlated to impairment of virus replication and mRNA expression. Applied Microbiology and Biotechnology, 97 12: 5245-5257. doi:10.1007/s00253-013-4835-8


Author Huynh, Hoai T.
Tran, Trinh T. B.
Chan, Leslie C. L.
Nielsen, Lars K.
Reid, Steven
Title Decline in baculovirus-expressed recombinant protein production with increasing cell density is strongly correlated to impairment of virus replication and mRNA expression
Journal name Applied Microbiology and Biotechnology   Check publisher's open access policy
ISSN 0175-7598
1432-0614
Publication date 2013-06-01
Year available 2013
Sub-type Article (original research)
DOI 10.1007/s00253-013-4835-8
Open Access Status Not Open Access
Volume 97
Issue 12
Start page 5245
End page 5257
Total pages 13
Place of publication Heidelberg, Germany
Publisher Springer
Language eng
Formatted abstract
The cell density effect is a well-established constraint in the baculovirus-insect cell expression platform, in which cell-specific productivity declines with increasing cell density, hence limiting the maximum achievable volumetric yield of protein product. A deeper elucidation of this phenomenon is sought in this study, by tracking the peak production of viral DNA (vDNA), recombinant LacZ mRNA, and β-galactosidase (β-gal) protein, over a wide range of cell densities. Sf9 suspension cell cultures were propagated in Sf-900 III serum-free medium and synchronously infected with rAcMNPV at multiple infection cell densities (ICDs) of between 0.5 and 8 × 106 cells/mL. There was a strong negative linear correlation between the specific β-gal yield and the peak cell density (PCD) post-infection, but contrary to previous reports, the yield decline started at a lower PCD of around 1 × 106 cells/mL. Most interestingly, there also was a corresponding strong negative linear correlation between the specific vDNA or LacZ mRNA yield, and the PCD. Comparing the infections at the highest and lowest PCDs tested, the yield decline was most dramatic for β-gal protein (95 %) and LacZ mRNA (90 %), while it was more moderate for vDNA (50 %). These declines were significantly reduced but not completely arrested, when spent medium was replaced with fresh at the ICD. These findings suggest that protein yield deterioration with increasing cell density originated from limitations during upstream events such as virus gene replication or transcription, rather than during the translational phase. Such limitations may be largely nutritional, but a more complex mechanism may be implicated
Keyword Baculovirus
Cell density effect
mRNA expression
Viral DNA replication
Infected Insect Cells
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Australian Institute for Bioengineering and Nanotechnology Publications
Official 2014 Collection
 
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