Objectives

The overarching goal of the project is to develop a model-based approach for engineering the Gram-positive bacterium B. subtilis and to develop synthetic modules for the production of metabolites and proteins of interest.

BaSynthec will:

  • Design and generate bacterial cells of reduced complexity by a model-driven experimental strategy for iterative deletion of chromosome parts. Focus will be put on the following traits: low energy waste, global deregulation of gene expression, and cell growth control
  • Generate quantitative data sets using a few selected strains lacking potentially expensive processes, hereafter termed “simpler” cells, to model at a systems level how some basic biological processes functionally integrate.
  • “Plug-in” new modules, which include entire pathways with their genetic regulations and their deregulated overexpression variants, in the “simpler” cell (experimentally) and in its cognate models (computationally). The selected modules will be used for establishing the proof-of-principle of the assembly of artificial operon/gene-clusters to generate functional synthetic modules to be “plugged-in”. Two modules of high biotechnological relevance will be constructed: i) the vitamin B5 biosynthetic pathway encoded by 5 genes, and ii) the secretion machinery for the export of extra-cellular enzymes.
  • Monitor quantitatively the operation of the “plugged-in” modules within the simpler host cell in order to tune and optimize the module. Input signals will be compounds inducing specifically a few promoters, each controlling the expression of a different key element of the module (or pathway). The output expression of these elements and others will be monitored in real time using luorescent reporters.
  • Apply both the methods and the gained knowledge about module plug-in in a synthetic cell to unravel high-level bacterial functions relevant to biotechnology (i.e. growth rate control).
  • Improve the safety of strains. By constructing a minimal genome strain as the basis for a cell factory, we considerably reduce its ability to survive in the environment and the possibility for unwanted side effects that are inherent to any natural organism. If successful, BaSynthec would deliver a cell factory with considerably fewer genes and proteins that is streamlined for performance in a bioreactor but has lost most of its ability to deal with natural environmental conditions. Hence, the probability for accidental gene transfer from this (or any future biotech host constructed in this fashion) is further reduced, as is the probability of undesired interactions with the product, humans or the environment. Thus, we believe that this project could actually become a major step forward in bio-safety.