Histidine-containing phosphocarrier protein (HPr) is a general component of the bacterial phosphoenolpyruvate:sugar phosphotransferase system (PTS) involved in the phosphorylation-coupled transport of numerous sugars called PTS sugars. HPr mainly exists in a dephosphorylated form in the presence of PTS sugars in the medium, while its phosphorylation increases in the absence of PTS sugars. A recent study revealed that the dephosphorylated form of HPr binds and antagonizes the function of the antisigma factor Rsd. This anti-sigma factor sequesters the housekeeping sigma factor σ(70) to facilitate switching of the sigma subunit on RNA polymerase from σ(70) to the stress-responsive sigma factor σ(S) in stationary-phase cells. In this study, the structure of the complex of Rsd and HPr was determined at 2.1 Å resolution and revealed that the binding site for HPr on the surface of Rsd partly overlaps with that for σ(70). The localization of the phosphorylation site on HPr at the binding interface for Rsd explains why phosphorylation of HPr abolishes its binding to Rsd. The mutation of crucial residues involved in the HPr-Rsd interaction significantly influenced the competition between HPr and σ(70) for binding to Rsd both in vitro and in vivo. The results provide a structural basis for the linkage of global gene regulation to nutrient availability in the external environment.
Carbon catabolite repression is a regulatory mechanism to ensure sequential utilization of carbohydrates and is usually accomplished by repression of genes for the transport and metabolism of less preferred carbon compounds by a more preferred one. Although glucose and mannitol share the general components, enzyme I and HPr, of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) for their transport, glucose represses the transport and metabolism of mannitol in a manner dependent on the mannitol operon repressor MtlR in Escherichia coli. In a recent study, we identified the dephosphorylated form of HPr as a regulator determining the glucose preference over mannitol by interacting with and augmenting the repressor activity of MtlR in E. coli. Here, we determined the X-ray structure of the MtlR-HPr complex at 3.5 Å resolution to understand how phosphorylation of HPr impedes its interaction with MtlR. The phosphorylation site (His15) of HPr is located close to Glu108 and Glu140 of MtlR and phosphorylation at His15 causes electrostatic repulsion between the two proteins. Based on this structural insight and comparative sequence analyses, we suggest that the determination of the glucose preference over mannitol solely by the MtlR-HPr interaction is conserved within the Enterobacteriaceae family.
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