Donald W. Kupke

Volume changes accompanying interaction of ligands with nucleic acids.

Publisher Summary This chapter provides a brief theoretical and experimental description of current techniques for the measurement of volume changes. The chapter illustrates the applicability of this approach for the interaction of ligands with nucleic acids. Specific examples have been provided from data obtained in laboratories; these include ligands with different physical binding modes, the contribution of nucleic acid conformation, bent deoxyoligonucleotides, and the formation of oligomer duplexes with chemically attached ligands. There are several techniques that are currently in use for the measurement of the volume change, ∆V, for an association reaction. Model-dependent techniques are based on the dependence of the equilibrium constant on hydrostatic pressure; model-independent techniques include direct measurements by dilatometry and indirect measurements of the mass and density of reactants and products of a particular reaction. The ∆V value is obtained from the measurements of the mass and the equilibrium density of solutions before and after mixing. The masses are obtained with a microbalance, and the densities are obtained with a magnetic suspension densimeter or with an Anton Paar (Graz Austria) densimeter.

Sequential volume changes on a small sample by density.

A procedure is described for generating a sequence of volume changes by density on the same sample of a solution. The entire volume-change profile over the reactant concentration range of interest can be accomplished with 0.1-0.2 mumol of a protein in 100 microliters of sample. The calcium-binding proteins, calmodulin and skeletal troponin-C, were employed to determine the volume increases attending the sequential addition of calcium ions. Both volume-change profiles exhibited nonlinear increases over the first four equivalents of calcium ion added.