Exploring alternative expression systems for expression of heterologous proteins and metabolic pathways

Englaender, Jacob Alexander
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Linhardt, Robert J.
Koffas, Mattheos A. G.
Maxwell, Patrick H.
Dordick, Jonathan S.
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Our next goal was to use chromosomal integration for metabolic engineering. Multiple studies, from our laboratory and others, have found that modulatin of gene expression is key to pathway optimization. Generally, this has been achieved using promoters of varying strengths on plasmids of variable copy number. However, chromosomal integration offers advantages for metabolic engineering that could resilt in increased production of high-value chemical products. Here, we have shown, usnig a model five-gene pathway for the production of the purple pigment violacein from tryptophan, again that the location of the integration matters. We found that of the four genomic locations used, only one showed any level of violacein production, and it was surprisingly not the same location that resulted in increased levels of mCherry expression. Interestingly, when the one-gene TAL pathway, which converts phenylalanine to trans-cinnamic acid, was integrated into the same loci, very little difference in production was observed between the locations. These results indicate that optimization of production may not only be location dependent, but it may also depend on the genes being expressed.
August 2016
School of Science
Dept. of Biological Sciences
Rensselaer Polytechnic Institute, Troy, NY
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