Overview
Our artificial futile cycle is driven by the expression of two key enzymes, isocitrate lyase and malate synthase, in glyoxylate cycle. These two enzymes are encoded by
aceA and
aceB genes, respectively. Since mammalian cells lacks genes expressing glyoxylate shunt, we introduce these genes form
E. coli and assemble them in a constitutive construct composed of mammalian constitutive promoter, mitochondrial leader sequence, tag protein and terminator that will allow the expression of prokaryotic gene in eukaryotic cell. In addition to the constitutive system, we assemble an inducible construct which will allow tunable gene expression according to the concentration of fatty acid in the medium. We decided to introduce inducible system to prevent fatty acid deficiency in low concentration of plasma fatty acid and facilitate greater fatty acid uptake at a high circulating fatty acid levels. The inducible and constitutive system will be compared in terms of fatty acid uptake rate in a range concentration of fatty acid.
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