Illuminating and exploiting programmed O-methylation in trans-AT polyketide synthases

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There is an urgent need to produce novel derivatives of natural product-based medicines, such as antibiotics and anticancer agents, to overcome drug resistance. Moreover, many herbicides, insecticides, and fungicides, which play an essential role in the protection of food crops, are natural products and new derivatives with lower toxicity and greater efficacy are needed to feed the burgeoning global population. Development of novel methodologies for structural modification of natural products and enhancing their production levels are important applications of synthetic biology. Many important natural products are built by modular multienzyme assembly lines. These offer strong potential for rational bioengineering to create novel natural product derivatives. However, a lack of detailed molecular insight into the structure, mechanism, and substrate tolerance of these inherently mobile protein machines has limited progress in this endeavour. Thus, there is a need to develop novel structure-based approaches to elucidating the mechanism and substrate specificity determinants of specific components of such machinery. This project aims to develop an atomic-level understanding of how assembly line machinery appends methyl groups to specific oxygen atoms during assembly of an important class of natural products known as polyketides. To do this, we will develop and apply strategies for capturing intrinsically dynamic components of these machines in specific functional states, which will facilitate the determination of their molecular structures using a variety of methods, including nuclear magnetic resonance spectroscopy, carbene-footprinting mass spectrometry and cryo-electron microscopy. We will also investigate the substrate tolerance of key parts of the machinery using chemically synthesised probe molecules. The new knowledge gained will inform future efforts to bioengineer these assembly lines to produce novel natural product derivatives with altered pharmacokinetic and pharmacodynamic profiles. This has the potential to exploited by UK-based pharmaceutical, agrochemical and biotechnology companies actively engaged in the development of natural product-based consumer products, which will ultimately benefit to wider society.