Role of Electrostatic Interactions in Transient Encounter Complexes in Protein-Protein Association Investigated by Paramagnetic Relaxation Enhancement

摘要

Protein—protein association can be viewed as a two-step process comprising the initial formation of an encounter complex ensemble followed by rearrangement, along a two-dimensional energy landscape, to form the final well-defined stereospecific complex (Figure 1). Theoretical work suggests that electrostatic interactions play an important role in encounter complex formation, thereby enhancing molecular association by permitting a reduced dimensionality search until the stringent orientational requirements for specific association are met. Recently, intermolecular paramagnetic relaxation enhancement (PRE) has been used to directly visualize an ensemble of lowly populated, highly transient encounter complexes in rapid exchange with the stereospecific complex. In this exchange regime, the observed intermolecular PREs are weighted population averages of the PREs of the species present, and depending on paramagnetic center-proton distances, species with an occupancy as low as 1% can be detected. For three relatively weak complexes (K_D ～ 1 -20 μM) from the bacterial phosphotransferase system, the distribution of nonspecific encounter complexes appeared to be qualitatively correlated to the electrostatic surface potentials of the interacting proteins. In this paper, we extend our previous work on the complex of the N-terminal domain of enzyme 1 (EIN) and HPr to examine the ionic strength dependence of intermolecular PREs and provide direct experimental evidence that the interactions involved in the formation of shortlived encounter complexes are predominantly electrostatic in nature.