Potential Regulatory Role of Competitive Encounter Complexes in Paralogous Phosphotransferase Systems

There are two paralogous Escherichia coli phosphotransferase systems, one for sugar import (PTS^sugar) and one for nitrogen regulation (PTS^Ntr), that utilize proteins enzyme I^sugar (EI^sugar) and HPr, and enzyme I^Ntr (EI^Ntr) and NPr, respectively. The enzyme I proteins have similar folds, as do their substrates HPr and NPr, yet they show strict specificity for their cognate partner both in stereospecific protein-protein complex formation and in reversible phosphotransfer. Here, we investigate the mechanism of specific EI^Ntr:NPr complex formation by the study of transient encounter complexes. NMR paramagnetic relaxation enhancement experiments demonstrated transient encounter complexes of EI^Ntr not only with the expected partner, NPr, but also with the unexpected partner, HPr. HPr occupies transient sites on EI^Ntr but is unable to complete stereospecific complex formation. By occupying the non-productive transient sites, HPr promotes NPr transient interaction to productive sites closer to the stereospecific binding site and actually enhances specific complex formation between NPr and EI^Ntr. The cellular level of HPr is approximately 150 times higher than that of NPr. Thus, our finding suggests a potential mechanism for cross-regulation of enzyme activity through formation of competitive encounter complexes.