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bioRxiv · 10.1101/2025.11.10.687708

Nanoplasmonic SERS reveals previously uncharacterised indole derivative in E. coli metabolism

Vali, M.·Wyatt, E.·Abbott, K.·Lio, P.·Croft, C.·Krauss, T.·Khan, M.·Zarkan, A.·Baumberg, J.·Fusco, D.
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Abstract

Tryptophanase (TnaA) is a promiscuous enzyme which regulates the production of amino acid-derived metabolites, essential for bacterial communication. TnaA is known to convert tryptophan into indole, an important signalling molecule influencing a range of behaviours - from bacterial quorum sensing to urinary tract infections (UTI) - but other conversion routes remain obscure. Here, we show that nanoplasmonic surface-enhanced Raman spectroscopy (SERS) combined with targeted genetic subtraction enables the label-free, cellular characterisation and structural identification of TnaA-derived metabolites directly from Escherichia coli culture solutions, without isolation or purification. We systematically profile the SERS spectra of wild-type and tnaA gene knockout strains (E. coli BW25113 and the uropathogenic 536) supplemented with each of the twenty amino acids, uncovering a previously unreported TnaA-dependent metabolic signature, I*, whose Raman fingerprint does not match free indole or any common indole derivative characterised by mass spectrometry. Using isotopomer-labelled substrates together with structural reference compounds, we perform label-free structural deduction of I* directly from SERS spectra. We find I*s structure is consistent with an intact indole ring carrying a C3 substituent, likely produced indirectly from intracellular tryptophan. These findings introduce a novel indole metabolite, raise new hypotheses regarding the biological role of TnaA-mediated byproducts in bacterial signalling and virulence, and demonstrate that nanoplasmonic SERS provides a powerful framework for probing enzyme activity and bioactive metabolite production directly from living bacterial cultures with nanomolar sensitivity and structural specificity.

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Vali, M., Wyatt, E., Abbott, K., Lio, P., Croft, C., Krauss, T., Khan, M., Zarkan, A., Baumberg, J., Fusco, D.. 2025-11-11. Nanoplasmonic SERS reveals previously uncharacterised indole derivative in E. coli metabolism. https://doi.org/10.1101/2025.11.10.687708

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