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Metabolic control at the cytosol–mitochondria interface in different growth phases of CHO cells

Paper ID Volume ID Publish Year Pages File Format Full-Text
31532 44810 2014 13 PDF Available
Title
Metabolic control at the cytosol–mitochondria interface in different growth phases of CHO cells
Abstract

•Dynamics of metabolic fluxes and compartmental enzyme activities in CHO cells.•Correlating functional, spatial and temporal information reveals metabolic control.•Rate-limiting steps in glycolysis and TCA cycle could be identified.•Glycolytic channeling affects pyruvate availability for mitochondria.•Anaplerotic enzymes and transaminases control the carbon supply into the TCA cycle.

Metabolism at the cytosol–mitochondria interface and its regulation is of major importance particularly for efficient production of biopharmaceuticals in Chinese hamster ovary (CHO) cells but also in many diseases. We used a novel systems-oriented approach combining dynamic metabolic flux analysis and determination of compartmental enzyme activities to obtain systems level information with functional, spatial and temporal resolution. Integrating these multiple levels of information, we were able to investigate the interaction of glycolysis and TCA cycle and its metabolic control. We characterized metabolic phases in CHO batch cultivation and assessed metabolic efficiency extending the concept of metabolic ratios. Comparing in situ enzyme activities including their compartmental localization with in vivo metabolic fluxes, we were able to identify limiting steps in glycolysis and TCA cycle. Our data point to a significant contribution of substrate channeling to glycolytic regulation. We show how glycolytic channeling heavily affects the availability of pyruvate for the mitochondria. Finally, we show that the activities of transaminases and anaplerotic enzymes are tailored to permit a balanced supply of pyruvate and oxaloacetate to the TCA cycle in the respective metabolic states. We demonstrate that knowledge about metabolic control can be gained by correlating in vivo metabolic flux dynamics with time and space resolved in situ enzyme activities.

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Keywords
Mammalian cell culture; CHO; Compartmentation; Enzyme activities; Metabolic flux analysis; Mitochondria; Metabolic switches
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Metabolic control at the cytosol–mitochondria interface in different growth phases of CHO cells
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Publisher
Database: Elsevier - ScienceDirect
Journal: Metabolic Engineering - Volume 23, May 2014, Pages 9–21
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us