Analysis of Transcription Factor Interactions at Sedimentation Equilibrium

Abstract

Publisher Summary This chapter explores the large numbers of proteins participating in the assemblies that regulate and catalyze transcription. Among methods available for characterizing their interactions, sedimentation equilibrium (SE) ultracentrifugation stands out as a direct and rigorous means of determining molecular masses, interaction stoichiometries, association constants, and the influences of low molecular weight effectors, ions, and crowding on the stabilities of protein complexes. The chapter also reviews the availability of modern instrumentation and the development of improved analysis methods that have resulted in an upsurge of interest in SE during the past decade. This chapter describes the application of SE techniques to the characterization of transcription factors and their interactions. It focuses on three situations that are encountered most frequently in studies of transcription factors: self-association, heteroassociation, and the presence of inactive components. Analysis of such interactions provides crucial information on the role of protein–protein interactions in the assembly of transcription and transcription–regulatory complexes.