Elucidation of a novel sialic acid metabolism pathway in mucus-foraging bacteria unravels mechanisms of adaptation to the gut

Andrew Bell, Jason Brunt, Emmanuelle Crost, Laura Vaux, Ridvan Nepravishta, David Owen, Dimitrios Latousakis, An Xiao, Wanqing Li, Xi Chen, Martin A. Walsh, Jan Claesen, Jesus Angulo, Gavin H. Thomas, Nathalie Juge (Lead Author)

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N-acetylneuraminic acid (Neu5Ac), the most abundant sialic acid form in humans, is commonly found in a terminal location on colonic mucins glycans where it is a much-coveted source of nutrients for gut bacteria. The mucin-foraging strategy of the human gut symbiont Ruminococcus gnavus is associated with the expression of an intramolecular trans-sialidase (IT-sialidase) that targets and cleaves off terminal α2–3 -linked Neu5Ac from glycoproteins, releasing 2,7-anhydro-Neu5Ac instead of Neu5Ac. Using a combination of in silico, molecular, biochemical and structural approaches, we have unravelled a unique metabolic pathway leading to the transport and metabolism of 2,7-anhydro-Neu5Ac which is underpinned by the exquisite specificity of the sialic acid transporter. The substrate binding protein, which forms part of a sialic acid transporter (SAT2) in R. gnavus ATCC29149, is specific to 2,7-anhydro-Neu5Ac, as shown by fluorescence spectroscopy, isothermal titration calorimetry (ITC), and saturation transfer difference nuclear magnetic resonance spectroscopy (STD NMR). Once inside the cell, 2,7-anhydro-Neu5Ac is converted into Neu5Ac via a novel enzymatic reaction catalysed by an oxidoreductase, RgNanOx. Following this conversion, Neu5Ac is then catabolised into N-acetylmannosamine (ManNAc) and pyruvate via the action of a Neu5Ac specific aldolase that is structurally and biochemically typical of NanA-like enzymes, as shownby X-ray crystallography of RgNanA wild-type and site-directed active site mutant K167A in complex with Neu5Ac. We confirmed the importance of this metabolic pathway in vivo by generating a R. gnavus nan cluster deletion mutant that lost the ability to grow on sialylated substrates. We showed that in gnotobiotic mice colonised with R. gnavus wild-type and mutant strains, the fitness of the nan mutant was significantly impaired as compared to the wild-type strain with a reduced ability to colonise the mucus layer. Overall, our study revealed a novel sialic acid pathway in bacteria, which has significant implications for the spatial adaptation of mucin-foraging gut symbionts in health and disease.
Original languageEnglish
Pages (from-to)2393–2404
Number of pages12
JournalNature Microbiology
Early online date21 Oct 2019
Publication statusPublished - Dec 2019

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