Transcriptional and metabolic response of recombinant Escherichia coli to spatial dissolved oxygen tension gradients simulated in a scale-down system.

摘要: Escherichia coli, expressing recombinant green fluorescent protein (GFP), was subjected to dissolved oxygen tension (DOT) oscillations in a two-compartment system for simulating gradients that can occur large-scale bioreactors. Cells were continuously circulated between the anaerobic (0% DOT) and aerobic (10% vessels of scale-down mimic an overall circulation time 50 s, mean residence compartments 33 17 respectively. Transcription levels mixed acid fermentation genes (ldhA, poxB, frdD, ackA, adhE, pflD, fdhF), measured by quantitative RT-PCR, increased 1.5- over 6-fold under oscillatory DOT compared cultures (constant 10% DOT). In addition, transcription level fumB whereas it decreased sucA sucB, suggesting tricarboxylic cycle functioning as two open branches. Gene revealed cytrochrome bd, which has higher affinity but lower energy efficiency, preferred cytochrome bO3 cultures. Post-transcriptional processing limited heterologous production system, inferred from similar gfp 19% GFP concentration Simulated also affected glyoxylate shunt (aceA), global regulators metabolism (fnr, arcA, arcB), other relevant (luxS, sodA, fumA, sdhB). Transcriptional changes explained observed alterations stoichiometric kinetic parameters, ethanol organic acids. Differences observed, indicating E. coli respond very fast intermittent conditions. The transcriptional responses reported here are useful establishing rational scale-up criteria strain design strategies improved culture performance at large scales.