Using the noncanonical metallo-amino Acid [Cu(II)(2,2′-Bipyridin-5-yl)]-alanine to study the structures of proteins - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Journal of Physical Chemistry Letters Année : 2023

Using the noncanonical metallo-amino Acid [Cu(II)(2,2′-Bipyridin-5-yl)]-alanine to study the structures of proteins

Résumé

Genetic code expansion allows modification of the physical and chemical properties of proteins by the site-directed insertion of noncanonical amino acids. Here we exploit this technology for measuring nanometer-scale distances in proteins. (2,2′-Bipyridin-5-yl)alanine was incorporated into the green fluorescent protein (GFP) and used as an anchoring point for Cu(II) to create a spin-label. The incorporation of (2,2′-bipyridin-5-yl)alanine directly into the protein resulted in a high-affinity binding site for Cu(II) capable of outcompeting other binding positions in the protein. The resulting Cu(II)-spin label is very compact and not larger than a conventional amino acid. By using 94 GHz electron paramagnetic resonance (EPR) pulse dipolar spectroscopy we have been able to determine accurately the distance between two such spin-labels. Our measurements revealed that GFP dimers can adopt different quaternary conformations. The combination of spin-labeling using a paramagnetic nonconventional amino acid with high-frequency EPR techniques resulted in a sensitive method for studying the structures of proteins.
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hal-04297922 , version 1 (21-11-2023)

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Leandro C. Tabares, Davis T. Daniel, José Luis Vázquez-Ibar, Cyrille Kouklovsky, Valérie Alezra, et al.. Using the noncanonical metallo-amino Acid [Cu(II)(2,2′-Bipyridin-5-yl)]-alanine to study the structures of proteins. Journal of Physical Chemistry Letters, 2023, 14 (14), pp.3368-3375. ⟨10.1021/acs.jpclett.3c00196⟩. ⟨hal-04297922⟩
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