Molecular recognition by proteins is fundamental to almost every biological process, particularly the protein associations underlying cellular signal transduction. Understanding the basis for protein-protein interactions requires the full characterization of the thermodynamics of their association. Historically it has been virtually impossible to experimentally estimate changes in protein conformational entropy, a potentially important component of the free energy of protein association. However, nuclear magnetic resonance spectroscopy has emerged as a powerful tool for characterizing the dynamics of proteins. Here we employ changes in conformational dynamics as a proxy for corresponding changes in conformational entropy. We find that the change in internal dynamics of the protein calmodulin varies significantly on binding a variety of target domains. Surprisingly, the apparent change in the corresponding conformational entropy is linearly related to the change in the overall binding entropy. This indicates that changes in protein conformational entropy can contribute significantly to the free energy of protein-ligand association.

A variety of technical issues such as the effects of macromolecular tumbling and the influence of competing relaxation mechanisms have also been largely resolved (vide infra). These advances have positioned solution NMR spectroscopy to efficiently and comprehensively characterize the fast internal dynamics of proteins of significant size. This review seeks to provide a compact but reasonably complete description of the theoretical and technical foundation for solution NMR relaxation methods that are currently being brought to bear on fast sub-nanosecond protein side chain dynamics and to present a summary of current findings and their possible significance. A survey of basic observations about side chain dynamics derived from NMR-relaxation studies is presented along with several analyses meant to dispel commonly held but apparently inaccurate correlations between dynamics, structure and function. How dynamics can enter into fundamental thermodynamic and kinetic aspects of protein function is also reviewed and illustrated with intriguing results from several systems that point to a promising future for this area of inquiry.

The physical basis for high affinity interactions involving proteins is complex and potentially involves a range of energetic contributions. Among these are changes in protein conformational entropy, which cannot yet be reliably computed from molecular structures. We have recently employed changes in conformational dynamics as a proxy for changes in conformational entropy of calmodulin upon association with domains from regulated proteins. The apparent change in conformational entropy was linearly related to the overall binding entropy. This view warrants a more quantitative foundation. Here we calibrate an “entropy meter” employing an experimental dynamical proxy based on NMR relaxation and show that changes in the conformational entropy of calmodulin are a significant component of the energetics of binding. Furthermore, the distribution of motion at the interface between the target domain and calmodulin are surprisingly non-complementary. These observations promote modification of our understanding of the energetics of protein-ligand interactions.