Chapitre D'ouvrage Année : 2020

Chapter Six - Bacterial expression and purification of vertebrate odorant-binding proteins

Résumé

Vertebrate odorant-binding proteins (OBPs) are small soluble proteins abundantly secreted in the olfactory mucus of many animal species, including humans. Vertebrate OBPs reversibly bind odorant molecules with micromolar range affinities. Although their physiological role is not clearly understood, OBPs are proposed to carry airborne odorants toward membrane olfactory receptors through the nasal mucus. Measurements of odorant-OBP interactions and structural studies require a large amount of pure OBPs devoid of ligands. The bacterial expression system is the first choice for expressing vertebrate OBPs used in our laboratory and others. This system generally produces OBPs in large amounts without major problems. In this chapter, we describe the milligram-scale production of recombinant pig OBP1 (pOBP1) in E. coli. The different steps of expression and purification are presented and discussed. Protocols for secondary structures investigation by circular dichroism and binding properties of the recombinant protein are also provided. More generally, these approaches can be used to produce and characterize any vertebrate OBPs for use in functional and structural studies.

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Dates et versions

hal-02917357 , version 1 (19-08-2020)

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Marine Brulé, Margot Glaz, Christine Belloir, Nicolas Poirier, Lucie Moitrier, et al.. Chapter Six - Bacterial expression and purification of vertebrate odorant-binding proteins. Methods in Enzymology, 642, Elsevier Inc., pp.125-150, 2020, ⟨10.1016/bs.mie.2020.05.002⟩. ⟨hal-02917357⟩
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