Genome synthesis of a universal synthetic host for antimicrobial drug production - towards the first deep-engineering of an actinobacterial genome

Antimicrobial resistance is an urgent problem that needs to be tackled by discovering new antimicrobial drugs. Genome sequencing has shown that microbial genomes encode the capability to produce an enormous variety of different chemical compounds, including thousands of potential antimicrobials. However, only a tiny fraction of these is currently being clinically used and the vast majority remains uncharacterised. A major bottleneck is the fact that the genetic machinery for compound production tends to be inactive under normal conditions - discovering and characterising antimicrobial leads usually requires their transfer to a heterologous host species. However, typical host candidates are limited in their ability to produce novel chemicals using the transferred genetic machinery from unrelated organisms.

In this project we will overcome this barrier using the powers of synthetic genomics. We will explore the common design features of the genomes of a particularly talented group of antimicrobial producers, in the class Actinobacteria, which have evolved a highly flexible metabolism pre-adapted to the production of high levels of a wide variety of compounds using horizontally acquired biosynthetic machinery. We will use this information and insights from earlier attempts to engineer antimicrobial high-production strains, to design a universal antibacterial host genome, which we will synthesise using an innovative combination of de-novo and template-based strategies, exploiting emerging genome synthesis techniques and advances in workflow automation. Creating an actinobacterial-derived universal synthetic host genome, which can produce a broad range of new chemicals compounds in a flexible, modular plug-and-play manner, will greatly expand our ability to access the antimicrobial treasures revealed by genome sequences. It will also facilitate the subsequent steps of compound modification and diversification using combinatorial approaches to create libraries of biosynthetic pathway variants. At the same time, the project work will greatly enhance the synthetic genomics capabilities in both host countries. Genome synthesis is still immature in many senses including expensive, labour-intensive, and not sufficiently automated. Here, we bring together complementary expertise to establish an innovative genome synthesis workflow suitable for tackling the major technical challenges of creating synthetic actinobacterial genomes.

These aims will be realised together by the strongly complementary teams of researchers from the University of Manchester and the Tokyo Institute of Technology and Nagoya University, in the UK and Japan, respectively. Together, our teams have expertise in the use of emerging technology in computational analysis (for redesigning the genome for synthesis), artificial intelligence/machine learning (to turn data into genome design strategies), automation (for creating synthetic genomes and DNA constructs), natural product research (to determine how the antimicrobials are produced and modified), microfluidics (for the rapid synthesis of genome parts in tiny volumes), and DNA chemistry (for designing new chemical reactions of DNA synthesis). Close interactions of all team members will be enabled by exchange visits to both the UK and Japan, during the project, as well as by organising an international symposium to bring together the natural products and engineering biology community.

Early career researchers and technicians are important members of our teams and will actively participate to the project, which will allow them to gain international experience and independence, while acquiring novel technical skills through the intense exchange visits.

The ActiSynth project will contribute to taking synthetic genomics to the next level. Its ambition is to establish a genome-synthesis pipeline that not only allows the genomes of model organisms to be synthesised, but that can in future be routinely used for metabolic engineering in challenging non-model species. We will use the GC-rich Actinobacteria as our proof-of-concept group, as these microbes combine challenging genomic complexity with biotechnological relevance as hosts for antimicrobial drug discovery and production. We will use comparative genomics and the UK partners' insights from earlier attempts to engineer antimicrobial production strains to design a universal antibacterial host genome, which we will begin to synthesise using an innovative combination of de-novo and template-based strategies, exploring emerging genome synthesis techniques and the development of a unique microfluidics-based synthesis workflow automation based on the approaches pioneered by the Japanese partners. Creating an actinobacterial-derived universal synthetic host genome, which can produce a broad range of new chemicals in a flexible, modular plug-and-play manner, will expand our ability to access the chemical diversity encoded by microbial genomes. It will also facilitate subsequent steps of compound modification and diversification using combinatorial approaches.
Realising these aims will depend on complementary expertise at the University of Manchester, the Tokyo Institute of Technology and Nagoya University, using emerging technology in computational analysis (for redesigning the genome for synthesis), artificial intelligence (to turn data into genome design strategies), automation (for creating synthetic genomes and DNA constructs), to natural product research (to determine how the antimicrobials are produced and modified), microfluidics (for rapid synthesis of genome parts in nanoliter volumes) and DNA chemistry (for designing new chemical reactions for DNA synthesis).