title: Probing structural transitions in both structured and disordered proteins using site-directed spin-labeling EPR spectroscopy.
abstract: EPR spectroscopy is a technique that specifically detects unpaired electrons. EPR-sensitive reporter groups (spin labels or spin probes) can be introduced into biological systems via site-directed spin-labeling (SDSL). The basic strategy of SDSL involves the introduction of a paramagnetic group at a selected protein site. This is usually accomplished by cysteine-substitution mutagenesis, followed by covalent modification of the unique sulfydryl group with a selective reagent bearing a nitroxide radical. In this review we briefly describe the theoretical principles of this well-established approach and illustrate how we successfully applied it to investigate structural transitions in both human pancreatic lipase (HPL), a protein with a well-defined α/β hydrolase fold, and the intrinsically disordered C-terminal domain of the measles virus nucleoprotein (N(TAIL) ) upon addition of ligands and/or protein partners. In both cases, SDSL EPR spectroscopy allowed us to document protein conformational changes at the residue level. The studies herein summarized show that this approach is not only particularly well-suited to study IDPs that inherently escape atomistic description by X-ray crystallography but also provides dynamic information on structural transitions occurring within well-characterized structured proteins for which X-ray crystallography can only provide snapshots of the initial and final stages.
authors.affiliation: Architecture et Fonction des Macromolécules Biologiques, UMR 6098 CNRS and Aix-Marseille Université, 163 Avenue de Luminy, Case 932, 13288 Marseille Cedex 09, France. Sonia.Longhi@afmb.univ-mrs.fr
authors.forename: Sonia
authors.initials: S
authors.lastname: Longhi
authors.forename: Valérie
authors.initials: V
authors.lastname: Belle
authors.forename: André
authors.initials: A
authors.lastname: Fournel
authors.forename: Bruno
authors.initials: B
authors.lastname: Guigliarelli
authors.forename: Frédéric
authors.initials: F
authors.lastname: Carrière
mesh_terms: D018360:Crystallography, X-Ray; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D006801:Humans; D008049:Lipase / Q000737:chemistry; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry*
chemical_list: D011506:Proteins; D008049:Lipase; C518354:PNLIP protein, human