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Tracking ligand-binding effect on protein stability by CD spectroscopy

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Abstract

The influence of protein-ligand interactions on protein stability is usually assessed by measurements in the liquid phase. CD spectroscopy appears to be a tool of choice to i) measure the conformation of the protein in different phases and to ii) follow the conformational changes of the protein upon binding. We therefore studied the stability of the rat odorant binding protein 3 (OBP3), its ability to remain functional and a preliminary test of its ligand binding specificity in the dry state. Solid-state spectra were performed on dry thin films prepared by drop casting of initial buffered aqueous solutions of the rat OBP3 onto optically transparent CaF2 windows and subsequently dried under low vacuum. We successfully recorded CD spectra of solid-state apo OBP3 from 280 to 130 nm. A previously unknown positive dichroic band became measurable in the solvent-free state at 175 nm. The reproducibility of the solid-state CD spectrum of apo OBP3 was confirmed by measuring several individually prepared films. We then assessed the time-dependent alteration of the protein in this dry environment. No change in the spectra was observed (storage at constant humidity with binary saturated salt solution), highlighting the stability of the OBP films on a monthly basis. Our results revealed that protein folding is not affected during film formation and remains stable over long-time scales.
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Dates and versions

hal-03790159 , version 1 (28-09-2022)

Identifiers

  • HAL Id : hal-03790159 , version 1

Cite

Jérémie Topin, Nykola C Jones, Søren V Hoffmann, Loïc Briand, Cornelia Meinert. Tracking ligand-binding effect on protein stability by CD spectroscopy. 18. international conference on Chiroptical Spectroscopy (ICCS), Jul 2022, New York, NY, United States. ⟨hal-03790159⟩
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