Structural and Functional Studies of the HAMP Domain of EnvZ, an Osmosensing Transmembrane Histidine Kinase in Escherichia coli

Abstract

The HAMP domain plays an essential role in signal transduction not only in histidine kinase but also in a number of other signal-transducing receptor proteins. Here Histidine kinases are the major signal transducer in bacteria, playing critical roles in adaptation to external stresses. EnvZ from Escherichia coli is one of the most extensively studied histidine kinases in terms of the functional and structural aspects (1). EnvZ is a transmembrane histidine kinase located in the inner membrane and functions as an osmosensor. It consists of a periplasmic receptor domain; two transmembrane domains, TM1 and TM2; and a cytoplasmic domain. The cytoplasmic domain has the bifunctional histidine kinase activity of EnvZ and can be further divided into three functional domains: the linker domain, domain A, and domain B. Domain A and domain B form the catalytic core of bifunctional histidine kinase EnvZ, having both kinase and phosphatase functions (2). Biochemical and structural studies of both domain A and domain B revealed that domain A contains the autophosphorylation site of a conserved His 243 and that it is a dimerization domain of EnvZ forming a four-helical bundle, whereas domain B contains an ATP-binding pocket (2-4). Thus, domain A and domain B are generally termed DHp and CA domains, respectively (5).The linker domain of EnvZ connects TM2 and the cytoplasmic domain. Although it is not directly involved in the enzymatic activities of EnvZ, it is known that the linker domain plays an important role in signal transduction. This is supported by the fact that a number of point mutations within the linker domain of EnvZ caused perturbation of the proper response against osmotic stresses (6 -9). The analysis of signal-transducing proteins from histidine kinases, adenylyl cyclases, methylaccepting chemotaxis proteins, and phosphatases revealed that like EnvZ these proteins also contain linker-like domains connecting their transmembrane domains to the cytoplasmic domains (10, 11). Importantly secondary structure prediction of these domains showed that they share a highly conserved helix-turn-helix fold even though their amino acid sequences have low homology. Therefore, these domains are called HAMP domains because this common motif was shared with histidine kinases, adenylyl cyclases, methyl-accepting chemotaxis proteins, and phosphatases. Mutational studies of HAMP domain of BarA (12), Aer (13), or NarX (14) also demonstrated that the HAMP domains play a crucial role in their signal transduction, supporting the notion that the HAMP domains are required for the proper signal transduction of histidine kinases, adenylyl cyclases, methyl-accepting chemotaxis proteins, and phosphatases (10, 11).To elucidate the mechanism of the signal transduction, functional hybrid histidine kinases between EnvZ and chemosensor Tar have been constructed by fusing the receptor domain and HAMP domain of Tar to the cytoplasmic kinase domain of EnvZ (15). The binding of aspartate to the receptor of this hybrid histidine kinase, termed Taz, m...

Salt-driven Equilibrium between Two Conformations in the HAMP Domain from Natronomonas pharaonis

“…Evidence for a dynamic conformation being an intrinsic feature of the HAMP domain have already been described in literature (13,14). Moreover, the NMR model was obtained well below the living temperature of the hyperthermophile A. fulgidus, which could have trapped the HAMP structure in one of its most stable conformations.…”

“…Despite these clear evidences for defined structural elements, indications for an intrinsic fragility of the HAMP domain are obvious. For example, the HAMP domain from NpHtrII was suggested to be in a molten-globule-like state at low salt concentration (13), whereas that from EnvZ, the osmosensing histidine kinase from Escherichia coli, was shown to be unable to form a stable structure by itself (14).…”

Probing bacterial transmembrane histidine kinase receptor–ligand interactions with natural and synthetic molecules

“…1A). CqsS lacks a HAMP domain, which is important in signal transduction in many histidine kinases (32)(33)(34)(35)(36)(37)(38). Thus, interactions between CAI-1 and the final transmembrane domain suggest that this transmembrane domain serves not only in ligand binding but as a critical regulatory region for CqsS kinase activity.…”

Significant Enhanced Expression and Solubility of Human Proteins in Escherichia coli by Fusion with Protein S from Myxococcus xanthus

“…Protein S can be readily dissociated from myxospores in its soluble form with EDTA. Previously, we also observed that when OmpR, an E. coli transcription factor, is fused to the N-terminal domain (NTD) of protein S, not only does the expression of the OmpR protein improve but also its solubility dramatically increases, by more than 20-fold (9). Importantly, the protein S-OmpR fusion protein exhibits DNA binding almost identical to that of OmpR alone (19).…”