1. Simple purification and characterization of soluble and homogenous ABC-F translation factors from Enterococcus faecium
- Author
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Demay, Fanny, Hallier, Marc, Georgeault, Sylvie, Com, Emmanuelle, Cattoir, Vincent, Goude, Renan, Gillet, Reynald, Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Institut de recherche en santé, environnement et travail (Irset), Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), École des Hautes Études en Santé Publique [EHESP] (EHESP), ARN régulateurs bactériens et médecine (BRM), Université de Rennes (UR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Infrastructures en Biologie Santé et Agronomie, IBiSA, Fondation pour la Recherche Médicale, FRM, (FRM EQU202203014663), Conseil Régional de Bretagne, and Université de Rennes 1
- Subjects
ABC-F protein ,Antibiotic resistance ,SUMO ,[SDV]Life Sciences [q-bio] ,Heparin chromatography ,Ni-NTA chromatography - Abstract
International audience; The family of ATP-binding cassette F proteins (ABC-F) is mainly made up of cytosolic proteins involved in regulating protein synthesis, and they are often part of a mechanism that confers resistance to ribosome-targeting antibiotics. The existing literature has emphasized the difficulty of purifying these recombinant proteins because of their very low solubility and stability. Here, we describe a rapid and efficient three-step purification procedure that allows for the production of untagged ABC-F proteins from Enterococcus faecium in the heterologous host Escherichia coli. After four purified ABC-F proteins were produced using this protocol, their biological activities were validated by in vitro experiment. In conclusion, our study provides an invaluable tool for obtaining large amounts of untagged and soluble ABC-F proteins that can then be used for in vitro experiments. © 2023 Elsevier Inc.
- Published
- 2023