Regulation of antibiotic production in Actinobacteria: new perspectives from the post-genomic era

The antimicrobial activity of many of their natural products has brought prominence to the Streptomycetaceae, a family of Gram-positive bacteria that inhabit both soil and aquatic sediments. In the natural environment, antimicrobial compounds are likely to limit the growth of competitors, thereby offering a selective advantage to the producer, in particular when nutrients become limited and the developmental programme leading to spores commences. The study of the control of this secondary metabolism continues to offer insights into its integration with a complex lifecycle that takes multiple cues from the environment and primary metabolism. Such information can then be harnessed to devise laboratory screening conditions to discover compounds with new or improved clinical value. Here we provide an update of the review we published in NPR in 2011. Besides providing the essential background, we focus on recent developments in our understanding of the underlying regulatory networks, ecological triggers of natural product biosynthesis, contributions from comparative genomics and approaches to awaken the biosynthesis of otherwise silent or cryptic natural products. In addition, we highlight recent discoveries on the control of antibiotic production in other Actinobacteria, which have gained considerable attention since the start of the genomics revolution. New technologies that have the potential to produce a step change in our understanding of the regulation of secondary metabolism are also described

Gene mining of biosynthesis genes and biosynthetic manipulation of marine bacteria for the production of new antibiotic candidates

Natural product drug discovery has traditionally been the corner stone of medicine having provided cures to many of today’s most common diseases. In particular, antibiotics have revolutionised healthcare and extended human lifespan. However, since their introduction into the clinic, resistance to these drugs has arisen. With the number of new antibiotics being discovered in recent years declining, and fewer drugs making it past clinical trials, we have reached the point where antibiotic resistant infections have become common place and a serious threat to health and society. There is now an urgent requirement for the discovery of new antibiotics and in particular those with unexploited mode of action. This thesis details the different areas of natural product drug development from discovery through to analogue generation. In Chapter one, the history of natural products as therapeutics is explored with a particular focus on antibiotics and how resistance arises against these agents. It outlines why the discovery of new antibiotics is so important and new methods used to facilitate this search. Chapter two follows with the development of a screening platform for antibiotic induction, using the model Streptomyces; Streptomyces coleiolor M145. A variety of culture additives are explored for their ability to induce secondary metabolism production. Chapter three then details the sampling and identification of microbes from a pseudo-marine environment and their screening for their ability to produce secondary metabolites with antibiotic properties. The second half of this thesis centres on the non-ribosomal peptide echinomycin. Collaborators Aquapharm supplied the marine derived strain AQP-4895, capable of producing echinomycin. Chapter four details the establishment of AQP-4895 culturing conditions and the shift observed in production profile. Next Chapter five looks at producing echinomycin analogues through precursor directed biosynthesis. A range of halogenated quinoxaline carboxylic acids are synthesised and fed to AQP-4895, and the respective echinomycin analogues monitored by LC-MS. Chapter Six then aims to direct biosynthesis of the halogenated analogues, using mutasynthesis. Due to the lack of genetic data available surrounding the strain, an unusual approach was taken, using iPCR to create a template for homologous recombination

The identification and heterologous expression of the biosynthetic gene cluster encoding the antibiotic and anticancer agent marinomycin

With the rise in antimicrobial resistance, there is an urgent need for new classes of antibiotic with which to treat infectious disease. Marinomycin, a polyene antibiotic from a marine microbe, has been shown capable of killing methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VREF), as well as having promising activity against melanoma. An attractive solution to the photoprotection of this antibiotic has been demonstrated. Here, we report the identification and analysis of the marinomycin biosynthetic gene cluster (BGC), and the biosynthetic assembly of the macrolide. The marinomycin BGC presents a challenge in heterologous expression due to its large size and high GC content, rendering the cluster prone to rearrangement. We demonstrate the transformation of Streptomyces lividans using a construct containing the cluster, and the heterologous expression of the encoded biosynthetic machinery and production of marinomycin B

Aromatic Polyketide GTRI-02 is a Previously Unidentified Product of the act

Microbial Biotechnolog

Large-Scale Bioinformatics Analysis of Bacillus Genomes Uncovers Conserved Roles of Natural Products in Bacterial Physiology

ABSTRACT Bacteria possess an amazing capacity to synthesize a diverse range of structurally complex, bioactive natural products known as specialized (or secondary) metabolites. Many of these specialized metabolites are used as clinical therapeutics, while others have important ecological roles in microbial communities. The biosynthetic gene clusters (BGCs) that generate these metabolites can be identified in bacterial genome sequences using their highly conserved genetic features. We analyzed an unprecedented 1,566 bacterial genomes from Bacillus species and identified nearly 20,000 BGCs. By comparing these BGCs to one another as well as a curated set of known specialized metabolite BGCs, we discovered that the majority of Bacillus natural products are comprised of a small set of highly conserved, well-distributed, known natural product compounds. Most of these metabolites have important roles influencing the physiology and development of Bacillus species. We identified, in addition to these characterized compounds, many unique, weakly conserved BGCs scattered across the genus that are predicted to encode unknown natural products. Many of these “singleton” BGCs appear to have been acquired via horizontal gene transfer. Based on this large-scale characterization of metabolite production in the Bacilli , we go on to connect the alkylpyrones, natural products that are highly conserved but previously biologically uncharacterized, to a role in Bacillus physiology: inhibiting spore development. IMPORTANCE Bacilli are capable of producing a diverse array of specialized metabolites, many of which have gained attention for their roles as signals that affect bacterial physiology and development. Up to this point, however, the Bacillus genus’s metabolic capacity has been underexplored. We undertook a deep genomic analysis of 1,566 Bacillus genomes to understand the full spectrum of metabolites that this bacterial group can make. We discovered that the majority of the specialized metabolites produced by Bacillus species are highly conserved, known compounds with important signaling roles in the physiology and development of this bacterium. Additionally, there is significant unique biosynthetic machinery distributed across the genus that might lead to new, unknown metabolites with diverse biological functions. Inspired by the findings of our genomic analysis, we speculate that the highly conserved alkylpyrones might have an important biological activity within this genus. We go on to validate this prediction by demonstrating that these natural products are developmental signals in Bacillus and act by inhibiting sporulation