Solution structure and dynamic character of the histidine-containing phosphotransfer domain of anaerobic sensor kinase ArcB from Escherichia coli

An Escherichia coli sensor kinase, ArcB, transfers a phosphoryl group to a partner response regulator in response to anaerobic conditions. Multidimens ional NMR techniques were applied to determine the solution structure of th e histidine-containing phosphotransfer signaling domain of ArcB (HPtArcB), which has a phosphorylation site, His717. The backbone dynamics were also i nvestigated by analyses of the N-15 relaxation data and amide hydrogen exch ange rates. Furthermore, the protonation states of the histidine imidazole rings were characterized by means of H-1 and N-15 chemical shifts at variou s pHs. The determined solution structure of HPtArcB contains five helices a nd forms a four-helix bundle motif like other HPt domains. The obtained ord er parameters, S-2, {H-1}-N-15 heteronuclear NOE values, and chemical excha nge parameters, R-ex, showed that the a-helical regions of HPtArcB are rigi d on both picosecond to nanosecond and microsecond to millisecond time scal es. On the other hand, helix D, which contains His717, exhibited low protec tion factors of less than 4000, indicating the presence of fluctuations on a slower time scale in helix D. These results suggest that HPtArcB may unde rgo a small conformational change in helix D upon phosphorylation. It was a lso shown that the imidazole ring of His717 has a pK(a) value of 6.76, whic h is similar to that of a solvent-exposed histidine imidazole ring, and tha t a pair of deprotonated neutral tautomers are rapidly exchanged with each other. This is consistent with the solution structure of HPtArcB, in which the imidazole ring of His717 is exposed to the solvent.