Monitoring of genes that respond to process-related stress in large-scale bioprocesses.
In large-scale aerobic fed-batch processes, cells are exposed to local zones of high glucose concentrations that can also cause local oxygen limitations at high cell densities. The mRNA levels of four stress genes (clpB, dnaK, uspA, and proU) and three genes responding to oxygen limitation or glucose excess (pfl, frd, and ackA) were investigated in an industrial 20-m(3) Escherichia coli process and in a scale-down reactor with defined high-glucose and low-oxygen zones. The mRNA levels of ackA and proU were high during the batch growth phase, but declined drastically when glucose became limited, whereas the mRNA levels of the other stress genes were relatively constant throughout the process. In the industrial-scale reactor, the stress gene mRNA levels were, in most cases, highest in the middle part and at the top of the reactor, where the substrate was fed. Cells passing through the high glucose zone of the scale-down reactor had elevated mRNA levels for the oxygen limitation genes and had also elevated heat-shock gene mRNA levels. Both responses to stress occurred within seconds. The approach presented in this study offers a tool for monitoring process-related changes in the transcriptional regulation of genes.
Schweder
T
T
Institute of Microbiology and Molecular Biology, Ernst-Moritz-Arndt-University, D-17487 Greifswald, Germany.
Krüger
E
E
Xu
B
B
Jürgen
B
B
Blomsten
G
G
Enfors
S O
SO
Hecker
M
M
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism; D019149:Bioreactors; D017931:DNA Primers; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005947:Glucose / Q000172:deficiency / Q000494:pharmacology; D006360:Heat-Shock Proteins / Q000378:metabolism*; D010100:Oxygen / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism
D001426:Bacterial Proteins; D017931:DNA Primers; D006360:Heat-Shock Proteins; D012333:RNA, Messenger; D005947:Glucose; D010100:Oxygen