Exploiting photoactivatable diazirines for the study of natural product biosynthesis and their mechanisms of action
Lead Research Organisation:
University of Warwick
Department Name: School of Life Sciences
Abstract
Natural products have been essential to the discovery of a wealth of industrially important drugs and agrochemicals. However, the rational design and development of drug leads requires the challenging identification of natural product targets and a fundamental understanding of their biosynthetic construction.
Diazirine-based photoactivatable probes provide a unique opportunity to investigate biological systems by capturing transient molecular interactions. Upon long wavelength UV (~360 nm) irradiation, diazirine moieties liberate nitrogen, forming highly reactive carbene species capable of C-H and heteroatom-H insertion and the generation of stable covalent crosslinks. Since the carbene intermediate is incredibly short lived, existing only for picoseconds, proximal chemical entities (e.g. amino acid residues) are rapidly labelled and can be mapped by mass spectrometry.
A multitude of biosynthetic pathways present attractive targets to exploit diazirine-based photoactivatable probes. By expanding current preparative methods of diazirines, this project aims to produce a library of photoactivatable substrates and biomolecules. For example, modification of a 4'-phosphopantetheine prosthetic which tethers intermediates to acyl carrier proteins (ACPs) in polyketide and fatty acid biosynthesis will allow mapping of inter-domain interactions and trajectory of the ACP.
This strategy will also be applied for selective incorporation of diazirine moieties into natural products, allowing downstream identification of their molecular targets using standard proteomics-based workflows. Here we will leverage the substrate promiscuity of catalytic domains responsible for selecting the starter and extender units in two PKS systems for which the molecular target is already established.
Taken together, this work will expand the biochemical applications and synthetic preparation of diazirines for the chemical biology toolkit.
Diazirine-based photoactivatable probes provide a unique opportunity to investigate biological systems by capturing transient molecular interactions. Upon long wavelength UV (~360 nm) irradiation, diazirine moieties liberate nitrogen, forming highly reactive carbene species capable of C-H and heteroatom-H insertion and the generation of stable covalent crosslinks. Since the carbene intermediate is incredibly short lived, existing only for picoseconds, proximal chemical entities (e.g. amino acid residues) are rapidly labelled and can be mapped by mass spectrometry.
A multitude of biosynthetic pathways present attractive targets to exploit diazirine-based photoactivatable probes. By expanding current preparative methods of diazirines, this project aims to produce a library of photoactivatable substrates and biomolecules. For example, modification of a 4'-phosphopantetheine prosthetic which tethers intermediates to acyl carrier proteins (ACPs) in polyketide and fatty acid biosynthesis will allow mapping of inter-domain interactions and trajectory of the ACP.
This strategy will also be applied for selective incorporation of diazirine moieties into natural products, allowing downstream identification of their molecular targets using standard proteomics-based workflows. Here we will leverage the substrate promiscuity of catalytic domains responsible for selecting the starter and extender units in two PKS systems for which the molecular target is already established.
Taken together, this work will expand the biochemical applications and synthetic preparation of diazirines for the chemical biology toolkit.
Organisations
People |
ORCID iD |
Studentship Projects
| Project Reference | Relationship | Related To | Start | End | Student Name |
|---|---|---|---|---|---|
| BB/T00746X/1 | 30/09/2020 | 29/09/2028 | |||
| 2887572 | Studentship | BB/T00746X/1 | 01/10/2023 | 30/09/2027 |