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  Metabolic adaptation of MDCK cells to different growth conditions: Effects on catalytic activities of central metabolic enzymes

Janke, R., Genzel, Y., Händel, N., Wahl, A., & Reichl, U. (2011). Metabolic adaptation of MDCK cells to different growth conditions: Effects on catalytic activities of central metabolic enzymes. Biotechnology and Bioengineering, 108(11), 2691-2704. doi:10.1002/bit.23215.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0013-8D7B-B Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0014-A380-D
Genre: Journal Article

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 Creators:
Janke, R.1, Author              
Genzel, Y.1, Author              
Händel, N.1, Author              
Wahl, A.1, Author              
Reichl, U.1, 2, Author              
Affiliations:
1Bioprocess Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society, ou_1738140              
2Otto-von-Guericke-Universität Magdeburg, ou_1738156              

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Free keywords: MDCK; enzymes; metabolism; glycolysis; glutaminolysis; pyruvate; metabolic adaptation
 Abstract: Lactate and ammonia are the most important waste products of central carbon metabolism in mammalian cell cultures. In particular during batch and fed-batch cultivations these toxic by-products are excreted into the medium in large amounts, and not only affect cell viability and productivity but often also prevent growth to high cell densities. The most promising approach to overcome such a metabolic imbalance is the replacement of one or several components in the culture medium. It has been previously shown that pyruvate can be substituted for glutamine in cultures of adherent Madin–Darby canine kidney (MDCK) cells. As a consequence, the cells not only released no ammonia but glucose consumption and lactate production were also reduced significantly. In this work, the impact of media changes on glucose and glutamine metabolism was further elucidated by using a high-throughput platform for enzyme activity measurements of mammalian cells. Adherent MDCK cells were grown to stationary and exponential phase in six-well plates in serum-containing GMEM supplemented with glutamine or pyruvate. A total number of 28 key metabolic enzyme activities of cell extracts were analyzed. The overall activity of the pentose phosphate pathway was up-regulated during exponential cell growth in pyruvate-containing medium suggesting that more glucose-6-phosphate was channeled into the oxidative branch. Furthermore, the anaplerotic enzymes pyruvate carboxylase and pyruvate dehydrogenase showed higher cell specific activities with pyruvate. An increase in cell specific activity was also found for NAD+-dependent isocitrate dehydrogenase, glutamate dehydrogenase, and glutamine synthetase in MDCK cells grown with pyruvate. It can be assumed that the increase in enzyme activities was required to compensate for the energy demand and to replenish the glutamine pool. On the other hand, the activities of glutaminolytic enzymes (e.g., alanine and aspartate transaminase) were decreased in cells grown with pyruvate, which seems to be related to a decreased glutamine metabolism. Copyright © 1999–2011 John Wiley & Sons, Inc. All Rights Reserved. [accessed July 29th, 2011]

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Language(s): eng - English
 Dates: 2011
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: Peer
 Identifiers: eDoc: 570618
Other: 16/11
DOI: 10.1002/bit.23215
 Degree: -

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Title: Biotechnology and Bioengineering
  Other : Biotechnol. Bioeng.
Source Genre: Journal
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Publ. Info: New York : Wiley [etc.]
Pages: - Volume / Issue: 108 (11) Sequence Number: - Start / End Page: 2691 - 2704 Identifier: ISSN: 0006-3592
CoNE: /journals/resource/111088195273104