Abstract
Publisher Summary This chapter presents the mathematical formalism to describe heterogeneous equilibria, with particular attention to the relationship between the macroscopic and microscopic equilibrium constants. Identification and characterization of critical macromolecular interactions within the cell are central research problems of the postgenomic era. Quantitative methods capable of accurately defining the stoichiometry, affinity, cooperativity, and thermodynamics are required for a fundamental mechanistic understanding and to effectively target molecular interactions for therapeutic intervention. Although many biologically significant interactions involve association of identical subunits, a much larger class of binding events involves interactions of dissimilar partners. Rigorous investigation of heterogeneous interactions presents particular challenges for experimental design, data analysis, and interpretation. Heterogeneous interactions involve at least two distinct macromolecular components. The features of several biophysical methods that are commonly used to characterize heterogeneous interactions are compared. Equilibrium and velocity analytical ultracentrifugation are particularly useful as baseline methods to define assembly models and extract equilibrium parameters. The focus in this chapter is on sedimentation equilibrium and this approach is illustrated with an analysis of a nonspecific protein–nucleic acid interaction.
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