Elucidating and exploiting molecular mechanisms of bacterial natural product biosynthesis

Project Outline

Natural product-based therapeutics are an essential part of modern medicine, and are also used widely in agriculture, e.g. as fungicides. However, resistance is neutralising their effectiveness. There is thus an urgent need to develop new natural product-based drugs and pesticides to overcome threats to health and sustainable agriculture posed by resistance. Most antimicrobials used in medicine and agriculture and many cancer chemotherapies derive from microbial natural products, which are often challenging to produce or modify by chemical synthesis. Our research program focuses on three key themes: 1) the discovery of novel antimicrobials and anticancer agents from diverse microorganisms, 2) elucidating the molecular mechanisms underlying their assembly, and 3) biosynthetic engineering to produce novel derivatives with enhanced therapeutic and agricultural potential. We employ a highly interdisciplinary approach, combining organic synthesis, analytical chemistry, microbiology, molecular genetics, genomics, bioinformatics, enzymology and structural biology, to address these problems.

In recent research, we have discovered antimicrobials with promising activity against Mycobacterium tuberculosis, which is responsible for millions of deaths every year in the developing world, Acinetobacter baumannii, identified by the World Health Organisation as one of three “critical priority” pathogens for new antibiotic research and development, and Pythium species which cause damping off disease in germinating crops. We have also discovered novel metabolites with potent and selective activity against cancer cell lines. Key steps in the biosynthesis of several antimicrobials, including gladiolin (active against multi-drug resistant M. tuberculosis), enacyloxin IIa (active against carbapenem-resistant A. baumannii), and bottromycin (active against methicillin-resistant Staphyloccocus aureus), and anticancer agents, such as gladiostatin and epoyketone proteasome inhibitors, have been elucidated. We have exploited the knowledge gained to create novel analogues of several of these metabolites, providing insights into their structure-activity relationships and producing potent derivatives that constitute a starting point for the development of new medicines and agrochemicals.

References

C. Huang, D. Zabala, E.L.C. de los Santos, L. Song, C. Corre, L.M. Alkhalaf and G.L. Challis. Parallelized gene cluster editing illuminates mechanisms of epoxyketone proteasome inhibitor biosynthesis. Nucleic Acids Res. 2023, 51, 1488-1499.

C. Hobson, M. Jenner, X. Jian, D. Griffiths, D.M. Roberts, M. Rey and G.L. Challis. Diene incorporation by a dehydratase domain variant in modular polyketide synthases. Nat. Chem. Biol. 2022, 18, 1410-1416.

S. Zhou, H. Bhukya, N. Malet, P. Harrison, D. Rea, M.J. Belousoff, H. Venugopal, P.K. Sydor, K.M. Styles, L. Song, M.J. Cryle, L.M. Alkhalaf, V. Fülöp, G.L. Challis and C. Corre. Molecular basis for control of antibiotic production by a bacterial hormone. Nature, 2021, 590, 463-467.

I.T. Nakou, M. Jenner, Y. Dashti, I. Romero-Canelón, J. Masschelein, E. Mahenthiralingam and G.L. Challis. Genomics-driven discovery of a novel glutarimide antibiotic from Burkholderia gladioli reveals an unusual polyketide synthase chain release mechanism. Angew. Chem. Int. Ed. 2020, 59, 23145-23153.

J. Masschelein, P.K. Sydor, C. Hobson, R. Howe, C. Jones, D.M. Roberts, Z.L. Yap., J. Parkhill, E. Mahenthiralingam and G.L. Challis. A dual transacylation mechanism for polyketide synthase chain release in enacyloxin antibiotic biosynthesis. Nat. Chem. 2019, 11, 906-912.

S. Kosol, A. Gallo, D. Griffiths, T.R. Valentic, J. Masschelein, M. Jenner, E.L.C de los Santos, L. Manzi, P.K. Sydor, D. Rea, S. Zhou, V. Fulop, N.J. Oldham, S.-C. Tsai, G.L. Challis and J.R. Lewandowski. Structural basis for chain release from the enacyloxin polyketide synthase. Nat. Chem. 2019, 11, 913-923.

A.J. Mullins, J.A.H. Murray, M.J. Bull, M. Jenner, C. Jones, G. Webster, T.R. Connor, J. Parkhill, G.L. Challis and E. Mahenthiralingam. Genome mining identifies cepacin as a key plant-protective metabolite of the biopesticidal bacterium Burkholderia ambifaria. Nat. Microbiol. 2019, 4, 996-1005.

Techniques

• Recombinant protein overproduction and purification
• Genetic manipulation
• Antimicrobial activity assays, including MIC determination
• Bioinformatics
• High resolution mass spectrometry of small molecules and proteins
• LC-MS analysis of microbial metabolites and enzymatic reaction products
• X-ray crystallographic analysis of proteins
• Semi-preparative HPLC purification of natural products and enzymatic reaction products
• Organic synthesis
• Small molecule structure elucidation using NMR and CD spectroscopy