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An integrated analysis of enzyme activities, cofactor pools and metabolic fluxes in baculovirus-infected Spodoptera frugiperda Sf9 cells

Paper ID Volume ID Publish Year Pages File Format Full-Text
23975 43486 2010 11 PDF Available
Title
An integrated analysis of enzyme activities, cofactor pools and metabolic fluxes in baculovirus-infected Spodoptera frugiperda Sf9 cells
Abstract

The scarcity of fundamental knowledge on the baculovirus–host cell interaction is a major drawback for the improvement of bioprocesses through Metabolic Engineering. After the first hours post-infection, the virus takes over the control of cellular machinery, leading to the repression of host gene expression and imposing a high metabolic burden to insect cells. Nevertheless, there is a lack of detailed data on the metabolic responses to infection, which are ultimately responsible for system productivity performance.In this work, a further insight into the central metabolism of Sf9 cells is achieved by a combined analysis of enzyme activities, cellular cofactors (ATP and NAD(P)+/NAD(P)H) and metabolic fluxes. Hexokinase and isocitrate dehydrogenase were identified as feasible limiting steps of metabolism; carbon and nitrogen metabolism enzymes were differentially regulated during batch cultures. Moreover, alterations occurring after infection demonstrated the importance of maintaining the energetic state of the cells for baculovirus replication, since ATP accumulated in a MOI-dependent way, and the glutamate dehydrogenase anaplerotic pathway was greatly activated. Altogether, cellular de-energization and stress responses are relevant factors in the metabolic burden imposed by infection. The implications for the improvement of bioprocess performance are discussed.

Keywords
AlaAT, Alanine aminotransferase; AspAT, Aspartate aminotransferase; CCI, Cell concentration at infection; EGFP, Enhanced green fluorescent protein; G6PDH, Glucose-6P dehydrogenase; GOGAT, Glutamate synthase (NADH-dependent); HexK, Hexokinase; ICDH, Isocit
First Page Preview
An integrated analysis of enzyme activities, cofactor pools and metabolic fluxes in baculovirus-infected Spodoptera frugiperda Sf9 cells
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 150, Issue 3, November 2010, Pages 332–342
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering