Chemical Analysis of Hybrid Fungal Megasynthases
Lead Research Organisation:
University of Bristol
Department Name: Chemistry
Abstract
Fungi produce a diverse array of biologically active compounds with roles as pharmaceuticals, agrochemicals and toxins. These include drugs such as the penicillins for treating bacterial infections, anticholesterol compounds such as the statins, and psychoactive drugs such as xenovulene. A major class of these compounds are the polyketides. In fungi polyketides are synthesised by giant multifunctional proteins known as polyketide synthases (PKS) - in turn these giant proteins are encoded by very large PKS genes. We have developed ways of isolating PKS genes from any given fungi which are involved in the biosynthesis of specific chemical compounds. For example we have found PKS gene clusters involved in the biosynthesis of the anticholesterol compound squalestatin, the mycotoxin fusarin A, the pigment tenellin and the psychoactive drug xenovulene from different fungal species. These PKS genes have been transferred to a new fungal host and switched on so that new chemical compounds are made. Despite very similar gene sequences, the four PKS genes encode proteins which can make very different chemical compounds. Thus, a knowledge of the gene sequence for these PKS does not yet help in understanding the processes which occur during the catalysis of polyketide formation in fungi. The PKS proteins evidently carry out a complex series of highly programmed chemical steps. While it is possible to read from the sequence the steps which could be catalysed, the programme controlling the order in which the steps are used, and the number of times they are repeated, is cryptic. The aim of this project is to find out how the giant PKS proteins are programmed. We will take a chemical approach to this problem. BBSRC and EU funded work is currently underway to generate numerous genetic changes in the genes themselves. We expect the genetic changes to lead to the production of modified proteins, which in turn should make new chemical compounds. By detecting, purifying and analysing the chemical structure of these new compounds we will be able to reveal the effect of the genetic change on the programme of the PKS. We expect to generate many tens of genetic experiments and each of these will require the examination of tens of genetic clones in fungi for the production of new compounds. We thus expect to have to examine hundreds to thousands of chemical extracts. We will use a state-of-the-art instrument to automate many of the purification and analysis steps. This instrument will combine High Performance Liquid Chromatography (HPLC) with sensitive, but robust, detectors which will be able to detect new compounds by size (evaporative light scattering detector), mass (mass spectrometer) and ultraviolet light (uv). The instrument will also be able to do automated purification for small samples, and will thus assist the chemist in processing the many hundreds or thousands of samples. We will also use the facilities of the School of Chemistry such as high field NMR and high resolution MS for structural elucidation. The new knowledge chemical structures of the new compounds produced from the mutant PKS will then allow us to elucidate the chemical effect of the genetic changes. We hope to eventually understand the link between gene sequence and chemical compound. This will allow two major advances - the ability to engineer fungal PKS at will to produce new compounds; and the ability to predict what compound will be made by simply reading a gene sequence.
Publications
Yakasai AA
(2011)
Nongenetic reprogramming of a fungal highly reducing polyketide synthase.
in Journal of the American Chemical Society
Wu J
(2008)
In vivo mutational analysis of the mupirocin gene cluster reveals labile points in the biosynthetic pathway: the "leaky hosepipe" mechanism.
in Chembiochem : a European journal of chemical biology
Williams K
(2016)
Heterologe Produktion pilzlicher Maleidride enthüllt die kryptische Cyclisierung in ihrer Biosynthese
in Angewandte Chemie
Williams K
(2017)
Genetic and chemical characterisation of the cornexistin pathway provides further insight into maleidride biosynthesis.
in Chemical communications (Cambridge, England)
Williams K
(2016)
Heterologous Production of Fungal Maleidrides Reveals the Cryptic Cyclization Involved in their Biosynthesis.
in Angewandte Chemie (International ed. in English)
Wasil Z
(2018)
Oryzines A & B, Maleidride Congeners from Aspergillus oryzae and Their Putative Biosynthesis.
in Journal of fungi (Basel, Switzerland)
Wasil Z
(2013)
One pathway, many compounds: heterologous expression of a fungal biosynthetic pathway reveals its intrinsic potential for diversity
in Chemical Science
Szwalbe AJ
(2019)
Characterisation of the biosynthetic pathway to agnestins A and B reveals the reductive route to chrysophanol in fungi.
in Chemical science
Szwalbe AJ
(2015)
Novel nonadride, heptadride and maleic acid metabolites from the byssochlamic acid producer Byssochlamys fulva IMI 40021 - an insight into the biosynthesis of maleidrides.
in Chemical communications (Cambridge, England)
Skellam EJ
(2010)
Mutation of key residues in the C-methyltransferase domain of a fungal highly reducing polyketide synthase.
in Molecular bioSystems
Description | The funding was used to buy analytical and preprative LCMS equipment which was used in research leading to > 18 publications. See the publication details attached. |
Exploitation Route | Details of all conclusions are contained in the publications, details of which are attached. |
Sectors | Chemicals Manufacturing including Industrial Biotechology Pharmaceuticals and Medical Biotechnology |
Description | BBSRC Grouped |
Amount | £4,519,651 (GBP) |
Funding ID | BB/K002341/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 01/2013 |
End | 12/2018 |
Description | BBSRC Grouped |
Amount | £857,230 (GBP) |
Funding ID | BB/I003355/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 07/2010 |
End | 07/2013 |
Description | BBSRC Grouped |
Amount | £4,519,651 (GBP) |
Funding ID | BB/K002341/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 01/2013 |
End | 12/2018 |
Description | BBSRC Grouped |
Amount | £857,230 (GBP) |
Funding ID | BB/I003355/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 07/2010 |
End | 07/2013 |
Description | BBSRC Grouped |
Amount | £558,465 (GBP) |
Funding ID | BB/J006289/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2012 |
End | 09/2015 |
Description | BBSRC Grouped |
Amount | £558,465 (GBP) |
Funding ID | BB/J006289/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 09/2012 |
End | 09/2015 |