Circular dichroism techniques for the analysis of intinsically disordered proteins and domains

Circular dichroism (CD) spectroscopy is a simple and powerful technique, whichallows for the assessment of the conformational properties of a protein or protein domain.Intrinsically disordered proteins (IDPs), as discussed throughout this series, differ fromrandom coil polypeptides in that different regions present specific conformationalpreferences, exhibiting dynamic secondary structure content (1). These dynamic secondarystructure elements can be stabilized or perturbed by different chemical (solvent, ionicstrength, pH) or physical (temperature) agents, by post-translational modifications, and byligands. This information is important for defining ID nature. As IDPs present dynamicconformations, circular dichroism measurements (and other approaches as well) should becarried out not as single spectra performed in unique conditions, but instead changing thechemical conditions and observing the behavior, as part of the determination of the ID nature.In this chapter, we present the basic methodology for performing Far-UV CDmeasurements on a protein of interest and for identifying and characterizing intrinsicallydisordered regions, and several protocols for the analysis of residual secondary structurepresent in the protein under study. These techniques are straightforward to perform; theyrequire minimal training and can be preliminary to more complex methodologies such asNMR.Fil: Chemes, Lucia Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Noval, María Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Noval, María Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: de Prat Gay, Gonzalo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin