Evaluation of Azido 3-Deoxy- d - Manno-oct-2-ulosonic Acid (Kdo) Analogues for Click Chemistry-Mediated Metabolic Labeling of Myxococcus xanthus DZ2 Lipopolysaccharide

Saïdi, Fares; Gamboa Marin, Oscar Javier; Veytia-Bucheli, José Ignacio; Vinogradov, Evgeny; Ravicoularamin, Gokulakrishnan; Jolivet, Nicolas Y.; Kezzo, Ahmad A.; Ramirez Esquivel, Eric; Panda, Adyasha; Sharma, Gaurav; Vincent, Stéphane P.; Gauthier, Charles ORCID: https://orcid.org/0000-0002-2475-2050 et Islam, Salim Timo ORCID: https://orcid.org/0000-0001-6853-8446 (2022). Evaluation of Azido 3-Deoxy- d - Manno-oct-2-ulosonic Acid (Kdo) Analogues for Click Chemistry-Mediated Metabolic Labeling of Myxococcus xanthus DZ2 Lipopolysaccharide ACS Omega , vol. 7 . pp. 34997-35013. DOI: 10.1021/acsomega.2c03711.

Prévisualisation

PDF - Version publiée Disponible sous licence Creative Commons Attribution. Télécharger (6MB) | Prévisualisation

Résumé

Metabolic labeling paired with click chemistry is a powerful approach for selectively imaging the surfaces of diverse bacteria. Herein, we explored the feasibility of labeling the lipopolysaccharide (LPS) of Myxococcus xanthusâÂ?Â?a Gram-negative predatory social bacterium known to display complex outer membrane (OM) dynamicsâÂ?Â?via growth in the presence of distinct azido (-N3) analogues of 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo). Determination of the LPS carbohydrate structure from strain DZ2 revealed the presence of one Kdo sugar in the core oligosaccharide, modified with phosphoethanolamine. The production of 8-azido-8-deoxy-Kdo (8-N3-Kdo) was then greatly improved over previous reports via optimization of the synthesis of its 5-azido-5-deoxy-d-arabinose precursor to yield gram amounts. The novel analogue 7-azido-7-deoxy-Kdo (7-N3-Kdo) was also synthesized, with both analogues capable of undergoing in vitro strain-promoted azide-alkyne cycloaddition (SPAAC) "click"chemistry reactions. Slower and faster growth of M. xanthus was displayed in the presence of 8-N3-Kdo and 7-N3-Kdo (respectively) compared to untreated cells, with differences also seen for single-cell gliding motility and type IV pilus-dependent swarm community expansion. While the surfaces of 8-N3-Kdo-grown cells were fluorescently labeled following treatment with dibenzocyclooctyne-linked fluorophores, the surfaces of 7-N3-Kdo-grown cells could not undergo fluorescent tagging. Activity analysis of the KdsB enzyme required to activate Kdo prior to its integration into nascent LPS molecules revealed that while 8-N3-Kdo is indeed a substrate of the enzyme, 7-N3-Kdo is not. Though a lack of M. xanthus cell aggregation was shown to expedite growth in liquid culture, 7-N3-Kdo-grown cells did not manifest differences in intrinsic clumping relative to untreated cells, suggesting that 7-N3-Kdo may instead be catabolized by the cells. Ultimately, these data provide important insights into the synthesis and cellular processing of valuable metabolic labels and establish a basis for the elucidation of fundamental principles of OM dynamism in live bacterial cells. ©

Type de document:	Article
Mots-clés libres:	-
Centre:	Centre INRS-Institut Armand Frappier
Date de dépôt:	03 févr. 2024 18:49
Dernière modification:	03 févr. 2024 18:49
URI:	https://espace.inrs.ca/id/eprint/13467

Gestion Actions (Identification requise)

Modifier la notice