Probing the heterologous metabolism supporting 6-deoxyerythronolide B biosynthesis in Escherichia coli.
Bottom Line: More specifically, the heterologous proteins responsible for 6dEB production were varied by adjusting their respective gene dosage levels.In this way, heterologous components required for posttranslational modification, 6dEB biosynthesis, and substrate provision were adjusted in expression levels to observe the relative effect each has on final heterologous biosynthesis.The results indicate that both the biosynthetic and substrate provision heterologous proteins impact 6dEB formation to a greater extent when compared with posttranslational modification and suggest these components for future protein and metabolic engineering.
Affiliation: Department of Chemical and Biological Engineering, Tufts University, Medford, MA 02155, USA.Show MeSH
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Mentions: To verify gene expression design, the strains used to vary heterologous gene dosage were first analysed by SDS‐PAGE. As seen in Fig. 1, protein levels correlate with gene dosage design. For example, the DEBS genes are observed when expressed from the pET expression plasmids but are not observed when expressed from low copy number within the chromosome. Figure 1 shows the same trend for the PCC and Sfp enzyme levels. Densitometry analysis was further used to assess and confirm expression differences within the SDS‐PAGE data.
Affiliation: Department of Chemical and Biological Engineering, Tufts University, Medford, MA 02155, USA.