Applications of Fluorescence Anisotropy in Understanding Protein Conformational Disorder and Aggregation

Abstract

Fluorescence spectroscopy is an ultra-sensitive multiparametric technique that provides key insights into protein conformational dynamics and size changes simultaneously. Fluorescence polarization (anisotropy) is one of the parameters related to the rotational dynamics of a fluorophore either intrinsic to the molecule or attached to a biomolecule. The anisotropy measurements can be utilized to unravel the structural and dynamical properties of biomolecules. The advantage of fluorescence anisotropy measurements is that it is a concentration-independent parameter; it can be measured either in the steady-state or in the time-resolved format. Steady-state fluorescence anisotropy provides important information about the overall size/dynamics of biomolecules, whereas the time-resolved fluorescence anisotropy can distinguish between the local and the global dynamics of a fluorophore. Therefore, the time-resolved anisotropy measurements allow one to determine the conformational flexibility as well as the size of biomolecules and assemblies. In recent years, it has been demonstrated that fluorescence anisotropy can be effectively utilized to obtain structural and dynamical information of protein-based assemblies such as aggregates, protein–lipid complexes etc. This chapter provides an overview of the applications of fluorescence anisotropy to study protein conformational disorder, misfolding and aggregation, leading to the formation of nanoscopic amyloid fibrils that are implicated in a range of human diseases.