Details
| Original language | English |
|---|---|
| Pages (from-to) | 602-613 |
| Number of pages | 12 |
| Journal | Biotechnology and bioengineering |
| Volume | 87 |
| Issue number | 5 |
| Publication status | Published - 17 Aug 2004 |
| Externally published | Yes |
Abstract
Guanosine-3′,5′-tetraphosphate (ppGpp) and σS, two regulators of the starvation response of Escherichia coli, have received increasing attention for monitoring cell physiological changes in production processes, although both are difficult to quantify. The kinetics of cAMP formation and degradation were not yet investigated in such processes, although the complex regulation of cAMP by synthesis, release, and degradation in connection with straightforward methods for analysis renders it a highly informative target. Therefore, we followed the cAMP concentration in various nonrecombinant and in four different recombinant glucose-limited fed-batch processes in different production scales. The intracellular cAMP concentration increases strongly at the end of the batch phase. Most cAMP is released to the cultivation medium. The rates of accumulation and degradation of extracellular cAMP are growth-rate-dependent and show a distinct maximum at a growth rate of about 0.35 h-1. At very low growth rates, below 0.05 h-1, extracellular cAMP is not produced but rather degraded, independent of whether this low growth rate is caused by glucose limitation or by the high metabolic load of recombinant protein production. In contrast to intracellular cAMP, which is highly unstable, analysis of extracellular cAMP is simpler and the kinetics of accumulation and degradation reflect well the physiological situation, including unlimited growth, limitation, and severe starvation of a production host.
Keywords
- cAMP, E. coli, Fed-batch cultivation, ppGpp, Recombinant protein production, RpoS
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biotechnology
- Chemical Engineering(all)
- Bioengineering
- Immunology and Microbiology(all)
- Applied Microbiology and Biotechnology
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In: Biotechnology and bioengineering, Vol. 87, No. 5, 17.08.2004, p. 602-613.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Change of extracellular cAMP concentration is a sensitive reporter for bacterial fitness in high-cell-density cultures of Escherichia coli
AU - Lin, Hongying
AU - Hoffmann, Frank
AU - Rozkov, Aleksei
AU - Enfors, Sven Olof
AU - Rinas, Ursula
AU - Neubauer, Peter
PY - 2004/8/17
Y1 - 2004/8/17
N2 - Guanosine-3′,5′-tetraphosphate (ppGpp) and σS, two regulators of the starvation response of Escherichia coli, have received increasing attention for monitoring cell physiological changes in production processes, although both are difficult to quantify. The kinetics of cAMP formation and degradation were not yet investigated in such processes, although the complex regulation of cAMP by synthesis, release, and degradation in connection with straightforward methods for analysis renders it a highly informative target. Therefore, we followed the cAMP concentration in various nonrecombinant and in four different recombinant glucose-limited fed-batch processes in different production scales. The intracellular cAMP concentration increases strongly at the end of the batch phase. Most cAMP is released to the cultivation medium. The rates of accumulation and degradation of extracellular cAMP are growth-rate-dependent and show a distinct maximum at a growth rate of about 0.35 h-1. At very low growth rates, below 0.05 h-1, extracellular cAMP is not produced but rather degraded, independent of whether this low growth rate is caused by glucose limitation or by the high metabolic load of recombinant protein production. In contrast to intracellular cAMP, which is highly unstable, analysis of extracellular cAMP is simpler and the kinetics of accumulation and degradation reflect well the physiological situation, including unlimited growth, limitation, and severe starvation of a production host.
AB - Guanosine-3′,5′-tetraphosphate (ppGpp) and σS, two regulators of the starvation response of Escherichia coli, have received increasing attention for monitoring cell physiological changes in production processes, although both are difficult to quantify. The kinetics of cAMP formation and degradation were not yet investigated in such processes, although the complex regulation of cAMP by synthesis, release, and degradation in connection with straightforward methods for analysis renders it a highly informative target. Therefore, we followed the cAMP concentration in various nonrecombinant and in four different recombinant glucose-limited fed-batch processes in different production scales. The intracellular cAMP concentration increases strongly at the end of the batch phase. Most cAMP is released to the cultivation medium. The rates of accumulation and degradation of extracellular cAMP are growth-rate-dependent and show a distinct maximum at a growth rate of about 0.35 h-1. At very low growth rates, below 0.05 h-1, extracellular cAMP is not produced but rather degraded, independent of whether this low growth rate is caused by glucose limitation or by the high metabolic load of recombinant protein production. In contrast to intracellular cAMP, which is highly unstable, analysis of extracellular cAMP is simpler and the kinetics of accumulation and degradation reflect well the physiological situation, including unlimited growth, limitation, and severe starvation of a production host.
KW - cAMP
KW - E. coli
KW - Fed-batch cultivation
KW - ppGpp
KW - Recombinant protein production
KW - RpoS
UR - http://www.scopus.com/inward/record.url?scp=4644340454&partnerID=8YFLogxK
U2 - 10.1002/bit.20152
DO - 10.1002/bit.20152
M3 - Article
C2 - 15352058
AN - SCOPUS:4644340454
VL - 87
SP - 602
EP - 613
JO - Biotechnology and bioengineering
JF - Biotechnology and bioengineering
SN - 0006-3592
IS - 5
ER -