Screening for new antibiotics using native protein mass spectrometry

Increasing antimicrobial resistance is one of the largest threats to our modern urbanised society. Amongst the multifaceted approach to tackling this problem is the need for new antibiotics.
Many new candidate antibiotics come from natural sources, such as microorganisms, where they are present as components of complex mixtures of other compounds. The challenge of identifying active compounds, often of low abundance, from such complex mixtures represents a serious bottleneck. This project aims to exploit a new technique developed by the Oldham group in which the complex mixture from a crude natural product extract is separated into its components using HPLC, and these combined in-line with a selected target protein. Binding is then directly measured by coupled native electrospray ionisation-mass spectrometry (ESI-MS). This approach promises to speed-up the
process of drug discovery by flagging-up interesting hit compounds within crude natural product extracts without the need for prior separation.

Native ESI-MS is an existing, specialist, technique able to detect non-covalent complexes between proteins and ligands. Separately, liquid chromatography (LC), especially high performance liquid chromatography (HPLC), is a commonly used for analysing and separating complex mixtures in a sample. LC is routinely coupled to MS, and commercial products are available. The novelty here is the combination of native MS with liquid chromatography. We have shown that the two are compatible, thus allowing complex natural product extracts to be interrogated for binding activity without preceding purification which is both challenging and time consuming.

The full PhD project will develop this platform for screening microbial extracts (provided by our collaborators at the University of Warwick) against antibiotic target proteins to detect
'hits' (compounds capable of binding to the target). This will facilitate screening of large numbers of potential antibiotic producing microorganisms for new compounds with activity against validated antibiotic targets.

Hits will be followed up by purification and full chemical/biological characterisation for potential medicinal use.

References to learn more:
1. Song H, et al. (1999) Crystal Structure of Intact Elongation Factor EF-Tu from Escherichia coli
in GDP Conformation at 2.05 A Resolution. J. Mol. Biol. 285: 1245-1256.
2. Kosol et al. (2019) Structural basis for chain release from the enacyloxin polyketide synthase.
Nat. Chem. 11: 913-923.