Regulation of polyhydroxyalkanoic acid synthase by riboswitch in Cupriavidus necator

Abstract

Riboswitches provide an opportunity for inducible gene expression at the transcriptional and translational levels. The riboswitches utilized in this thesis function by creating secondary mRNA structures that sequesters the ribosome binding site (RBS), preventing translation. This secondary structure can bind to theophylline, causing the mRNA to refold, exposing the RBS and allowing translation to occur. Riboswitches are currently underutilized, tested in only a few model organisms and primarily explored through reporter genes for proof-of-concept. This thesis demonstrates the functionality of theophylline riboswitches in the industrially relevant bacteria Cupriavidus necator, and for inducible control of the industrially relevant enzyme polyhydroxyalkanoic acid synthase (phaC), both of which have previously not been demonstrated. This thesis tests the applicability of the riboswitch in industry-relevant bacteria and addresses a significant gap in the knowledge about riboswitch functionality.