GEN2NCE - a synthetic biology platform for natural product discovery

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Microbial natural products and their synthetic derivatives are widely used in the pharmaceutical and agrochemical industries. Of all small-molecule new chemical entities (NCEs) that entered the market from 1981-2006, 34% were natural products or semisynthetic derivatives. In particular, natural products formed the basis for a large number of new antimicrobial (68 % of NCE), anticancer (54 % of NCE) and agrochemical (40% of NCE) agents. However, natural products have become de-prioritised as resource for the discovery of new pharmaceutical and agrochemical leads over the last two decades due to the high rate of rediscovery of known compounds using traditional bioactivity screening-based approaches.Over the past five years, genome-sequencing technologies have improved greatly and it is now possible to obtain a complete bacterial genome sequence for less than £500. Bioinformatics analyses of such genome sequences have revealed that many bacteria harbour the capability to assemble a far a greater number of potentially useful natural products than are typically observed in laboratory cultures. This is particularly true of Actinobacteria, which typically contain 20-50 biosynthetic pathways, but only produce a handful of the corresponding metabolic products. The underlying reason for this appears to be that the genes encoding such pathways are poorly expressed under laboratory growth conditions. Several methods for inducing the expression of these gene clusters have been reported in the literature, but few appear to be general. The development of general methods for the activation of silent biosynthetic gene clusters promises to yield a wealth of novel bioactive natural products with potential applications in medicine and agriculture.In 2011, we reported a method for activating silent biosynthetic gene clusters in Actinobacteria involving the constitutive expression of particular type of transcriptional activator gene. In follow up work funded by the BBSRC, we have sought to establish whether it has broad applicability. Through a strategic longer or larger award (sLoLa) we have shown that this type of activator gene is commonly associated with natural product biosynthetic gene clusters in Actinobacteria. Moreover, we have shown that constitutive expression of such activator genes is able to turn on various biosynthetic gene clusters that express poorly in laboratory cultures, leading to the discovery of several novel natural products. In other work carried out as part of the "engineering biosynthetic pathways" theme of the Warwick Integrative Synthetic Biology (WISB) Centre, we have developed efficient methods for rapid cloning of entire biosynthetic gene clusters. This opens the path for using our gene cluster activation technology in heterologous hosts.We have recently completed Innovate UK Innovation and Commercialisation of University Research and BBSRC Pathfinder projects, which have demonstrated a demand for novel structurally diverse natural products to feed agrochemical and pharmaceutical discovery pipelines. The stage is now set for development of our science into a commercial technology for the production of novel natural product libraries. To do this, however, we need to increase the throughput of our approach. This will facilitate the production of larger libraries that are suitable for use with high-throughput screens. We aim to develop heterologous hosts that allow co-expression of a biosynthetic gene cluster and an activator gene under the control of an inducible or growth-phase dependent promoter. We also aim increase the throughput of our gene cluster activation technology by employing robotics available through WISB to automate it. By the end of the project, we aim to have produced a compound library of sufficient size to supply to early adopters as a screening resource.