ASSEMBLY OF SYNTHETIC ORGANELLES IN BUDDING YEAST

Abstract

The spatial control of metabolism is an emerging trend in synthetic biology. Controlling the intracellular location of engineered reactions can endow microbial factories with improved or novel functionalities. The budding yeast, Saccharomyces cerevisiae, is a well-proven microbial chassis for housing synthetic bioprocesses. Establishing bioorthogonal organelles in yeast may ameliorate longstanding problems, such as undesired side reactions and toxicity, associated with such bioprocesses. In this thesis, two systems – bacterial microcompartments (MCP) and membraneless organelles (MO) – were chosen for recapitulation as synthetic organelles in yeast. A streamlined DNA assembly methodology was established for expression of these multi-component systems. The expression of MCPs and MOs in yeast was characterized by bioimaging methods. Further functional interrogation of both MCP and MO components reveals novel protein-protein interactions that underlie the plasticity behind their assembly. The work presented herein lays the groundwork for installing synthetic organelles in yeast, with implications for bioengineering of this versatile microorganism.