coli MG1655 Δ arcA Δ iclR and E. coli BL21 (DE3) cultivated under glucose abundant conditions. The ratios, shown in Figure 6, were used as constraints to determine net fluxes. Standard errors are calculated by propagating measured errors of extracellular fluxes and ratios. Absolute fluxes in were rescaled to the glucose uptake rate (shown in the upper boxes) to allow a clear https://www.selleckchem.com/products/iacs-010759-iacs-10759.html comparison in
flux distribution between the different strains. MK 8931 in vivo A possible hypothesis is the following. Microarray data and Northern blot analysis showed that genes coding for enzymes participating in reactions involving gluconeogenesis, the TCA cycle and glycogen biosynthesis were upregulated in E. coli BL21 compared to E. coli K12 . The higher aceA and aceB transcription in BL21 is caused by the apparent lower transcription of the iclR repressor . Consequently, lower IclR levels are present in the cell and the glyoxylate pathway is active . The lower transcription of iclR in E. coli BL21 may be explained by two mutations Captisol molecular weight in the iclR promoter region compared to E. coli K12 MG1655 (BLAST analysis, Figure 8). Particularly the mutation close to the Pribnow box or -10 box is important as it can have a major effect on the binding of RNA polymerase and hence gene
expression [62, 63]. Figure 8 BLAST analysis of the iclR promoter. Basic Local Alignment Search of the promoter region of iclR in an E. coli K12 MG1655 and BL21 reveals 2 mutations (highlighted by boxes) in the BL21 strain. The binding sites of the regulators FadR and IclR (autoregulator) are underlined. TS stands for transcription start. Results were obtained using the Interleukin-3 receptor NCBI online tool http://blast.ncbi.nlm.nih.gov. Not only is the glyoxylate flux similar, the TCA flux is improved as well in both strains compared to the E. coli K12 MG1655 wild type. Release of repression on transcription of TCA genes explains the higher flux in E. coli K12 ΔarcAΔiclR , and this must also be valid for E. coli BL21 as transcription of its TCA genes was highly upregulated compared to E. coli K12 . Genome comparison showed that although
BL21 and K12 genomes align for > 99%, many minor differences appear, which can explain the metabolic differences observed [64, 65]. However, those studies did not focus on differences in arcA regions. Using a Basic Local Alignment Search Tool (BLAST) it was determined that there is a 99% similarity in the arcA gene between the two strains. Only five minor mutations are observed (BLAST results shown in Additional file 3). However, the consequence of these mutations is that five other codons are formed in the mRNA in BL21 as opposed to MG1655 (see Table 4). These different codons in BL21 still encrypt for the same amino acids but two of these five codons (i.e. CUA and UCC) are known low-usage codons in E. coli and can cause translational problems [66, 67].