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Engineering Actinobacteria to enhance natural product biosynthesis

Principal supervisor: Dr Helen O'Hare, Department of Molecular and Cell Biology

Industry partner: Isomerase Therapeutics

Project Title: Engineering Actinobacteria to enhance natural product biosynthesis

University of registration: University of Leicester

Project outline:

The Actinobacteria are important producers of approved drugs (>$1 billion dollars in 2016) but low yields of most natural products are a significant barrier to drug discovery, development and commercialisation. New approaches to improve yield would benefit the biotechnology industry and human, animal and plant health.

The nutritional state of the bacteria is a critical component in switching on natural product biosynthesis. Previous studies have identified a metabolic regulator GarA that is important for glutamate production by industrial strains of Corynebacterium glutamicumand for growth of Mycobacterium tuberculosis. GarA regulates the tricarboxylic acid cycle by inhibiting enzyme activity, and the protein is conserved in antibiotic-producing Actinobacteria.

Engineering the GarA regulatory pathway could induce natural product gene clusters and improve the supply of metabolic precursors for biosynthesis.

The BBSRC Natural Products Discovery and Bioengineering Network funded a collaboration between the University of Leicester and Isomerase Therapeutics to carry out a Proof of Concept study in 2018. The results demonstrate the importance of GarA in industrial strains ofSaccaropolyspora erythraeaand the potential to increase the yield of erythromycin production by engineering this pathway: overexpression of GarA increased the yield of erythromycin by 30% in flask fermentation.

We propose to determine the mechanism by which GarA promotes erythromycin biosynthesis (the relative contributions of gene expression changes, changes in the phosphoproteome and changes in the metabolome and precursor supply). The student will investigate how best to engineer the pathway (chemical or genetic approaches) and determine whether the approach could be applied broadly to diverse Actinobacteria.

This project will address basic questions about the stimuli controlling the GarA pathway and antibiotic production in natural and industry-relevant environments. The industrial placement will be critical to assess the impact of engineering GarA on the metabolome and antibiotic-yield within the fermenters at Isomerase Therapeutics.

Objectives:

  1. Determine the effect of GarA overexpression on the metabolome and transcriptome of S. erythraeagrown in flask and fermenter: what is the mechanism by which GarA increases antibiotic production?
  2. Determine the dynamics of GarA expression and phosphorylation in natural and lab environments. What are the stimuli for expression of GarA and kinases, and what are the stimuli for kinase activity? (amino acids and redox status are important regulators of this pathway in related organisms)
  3. Investigate the potential of our panel or GarA variants to selectively regulate enzyme activity and metabolic flux.

Contact: Dr Helen O'Hare, University of Leicester