The Use of Heterologous Expression Platforms for the Production of Novel Actinomycete Antibiotics

Alford, Abigail (2021) The Use of Heterologous Expression Platforms for the Production of Novel Actinomycete Antibiotics. Doctoral thesis, University of East Anglia.

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Antimicrobial resistance (AMR) poses a considerable threat to modern living. To combat this threat methods for the discovery and diversification of antibiotics are of great importance. This project focuses on the use of heterologous expression for the rapid production and structural diversification of natural products (NPs). Firstly, I exemplify a synthetic heterologous platform for class II type B lantipeptide production. Further to this, I expand on the use of Saccharopolyspora erythraea ISOM as a heterologous expression strain for NPs.

Lantipeptides are Ribosomally-synthesised and Post-translationally modified Peptides (RiPPs) with clinical utility as antibacterials and treatments for cystic fibrosis. Rapid peptide diversification was achieved by Gibson assembly using a library of oligonucleotides encoding lantipeptide core peptides and an engineered precursor peptide gene (lanA) housed within a synthetic cassette. The production of 83 novel lantipeptides was achieved, with 61 showing potential as antibacterials. Seven lantipeptides were purified and their structure and bioactivity characterised.

Production of actinomycetes cryptic NPs in S. erythraea ISOM was mostly unsuccessful. Only production of fasamycin C was achieved using a de-repressed formicamycin biosynthetic gene cluster (BGC). The de-repressed formicamycin BGC was used to compare NP expression profiles within the parental strain and within different heterologous hosts. The heterologous hosts had significantly different expression profiles of the NPs compared to the parental strain. A further six new glycosylated fasamycin-derived compounds were produced from heterologous host S. erythraea ISOM, exemplifying how heterologous production may be used for the diversification of NPs.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Biological Sciences
Depositing User: Chris White
Date Deposited: 31 May 2022 13:58
Last Modified: 31 May 2022 13:58


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