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Development of RNA-based translation modulation tools for synthetic biology applications
(Lethbridge, Alta. : Universtiy of Lethbridge, Department of Mathematics and Computer Science, 2019) Hakstol, Rhys J.; University of Lethbridge. Faculty of Arts and Science; Wieden, Hans-Joachim
The ability to reliably regulate translation of heterologous proteins is of great interest for diverse applications in the field of biological and metabolic engineering. RNA-based tools to control ribosome-dependent synthesis of proteins within E. coli were developed. First, adaptation of the well-studied RNA-IN and RNA-OUT system based on E. coli Tn10 to reliably repress translation regardless of coding sequence context was performed. Using an invariant sequence within the RNA-IN/OUT interacting region does not interfere with the function of the RNAs involved. The same RNA-IN/RNA-OUT system was adapted to activate the translation of a specific target mRNA. Regulation of translation extends to processing of polyproteins. In eukaryotes this can be accomplished by viral 2A peptides, avoiding reinitiating at a downstream start codon but has not been observed in bacteria. Therefore, a strategy for developing libraries of 2A peptides with potential activity across an array of organisms was developed.