Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.13.3 (histidine kinase)
2,405 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The structure of the water-soluble, periplasmic domain of the fumarate sensor DcuS (DcuS-pd) has been determined by NMR spectroscopy in solution. DcuS is a prototype for a sensory histidine kinase with transmembrane signal transfer. DcuS belongs to the CitA family of sensors that are specific for sensing di- and tricarboxylates. The periplasmic domain is folded autonomously and shows helices at the N and the C terminus, suggesting direct linking or connection to helices in the two transmembrane regions. The structure constitutes a novel fold. The nearest structural neighbor is the Per-Arnt-Sim domain of the photoactive yellow protein that binds small molecules covalently. Residues Arg107, His110, and Arg147 are essential for fumarate sensing and are found clustered together. The structure constitutes the first periplasmic domain of a two component sensory system and is distinctly different from the aspartate sensory domain of the Tar chemotaxis sensor.
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PMID:The NMR structure of the sensory domain of the membranous two-component fumarate sensor (histidine protein kinase) DcuS of Escherichia coli. 1290 89

Histidine kinases are used extensively in prokaryotes to monitor and respond to changes in cellular and environmental conditions. In Bacillus subtilis, sporulation-specific gene expression is controlled by a histidine kinase phosphorelay that culminates in phosphorylation of the Spo0A transcription factor. Sda provides a developmental checkpoint by inhibiting this phosphorelay in response to DNA damage and replication defects. We show that Sda acts at the first step in the relay by inhibiting autophosphorylation of the histidine kinase KinA. The structure of Sda, which we determined using NMR, comprises a helical hairpin. A cluster of conserved residues on one face of the hairpin mediates an interaction between Sda and the KinA dimerization/phosphotransfer domain. This interaction stabilizes the KinA dimer, and the two proteins form a stable heterotetramer. The data indicate that Sda forms a molecular barricade that inhibits productive interaction between the catalytic and phosphotransfer domains of KinA.
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PMID:Structure and mechanism of action of Sda, an inhibitor of the histidine kinases that regulate initiation of sporulation in Bacillus subtilis. 1502 39

Overexpression of proteins in Escherichia coli at low temperature improves their solubility and stability. Here, we apply the unique features of the cspA gene to develop a series of expression vectors, termed pCold vectors, that drive the high expression of cloned genes upon induction by cold-shock. Several proteins were produced with very high yields, including E. coli EnvZ ATP-binding domain (EnvZ-B) and Xenopus laevis calmodulin (CaM). The pCold vector system can also be used to selectively enrich target proteins with isotopes to study their properties in cell lysates using NMR spectroscopy. We have cloned 38 genes from a range of prokaryotic and eukaryotic organisms into both pCold and pET14 (ref. 3) systems, and found that pCold vectors are highly complementary to the widely used pET vectors.
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PMID:Cold-shock induced high-yield protein production in Escherichia coli. 1522 43

Circadian oscillators are endogenous biological systems that generate the approximately 24 hour temporal pattern of biological processes and confer a reproductive fitness advantage to their hosts. The cyanobacterial clock is the simplest known and the only clock system for which structural information for core component proteins, in this case KaiA, KaiB and KaiC, is available. SasA, a clock-associated histidine kinase, is necessary for robustness of the circadian rhythm of gene expression and implicated in clock output. The N-terminal domain of SasA (N-SasA) interacts directly with KaiC and likely functions as the sensory domain controlling the SasA histidine kinase activity. N-SasA and KaiB share significant sequence similarity and, thus, it has been proposed that they would be structurally similar and may even compete for KaiC binding. Here, we report the NMR structure of N-SasA and show it to be different from that of KaiB. The structural comparisons provide no clear details to suggest competition of SasA and KaiB for KaiC binding. N-SasA adopts a canonical thioredoxin fold but lacks the catalytic cysteine residues. A patch of conserved, solvent-exposed residues is found near the canonical thioredoxin active site. We suggest that this surface is used by N-SasA for protein-protein interactions. Our analysis suggests that the structural differences between N-SasA and KaiB are the result of only a few critical amino acid substitutions.
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PMID:Structure of the N-terminal domain of the circadian clock-associated histidine kinase SasA. 1531 3

DcuS is a membrane-associated sensory histidine kinase of Escherichia coli specific for C(4) -dicarboxylates. The nature of the stimulus and its structural prerequisites were determined by measuring the induction of DcuS-dependent dcuB'-'lacZ gene expression. C(4)-dicarboxylates without or with substitutions at C2/C3 by hydrophilic (hydroxy, amino, or thiolate) groups stimulated gene expression in a similar way. When one carboxylate was replaced by sulfonate, methoxy, or nitro groups, only the latter (3-nitropropionate) was active. Thus, the ligand of DcuS has to carry two carboxylate or carboxylate/nitro groups 3.1-3.8 A apart from each other. The effector concentrations for half-maximal induction of dcuB'-'lacZ expression were 2-3 mm for the C(4)-dicarboxylates and 0.5 mm for 3-nitropropionate or d-tartrate. The periplasmic domain of DcuS contains a conserved cluster of positively charged or polar amino acid residues (Arg(107)-X(2)-His(110)-X(9)-Phe(120)-X(26)-Arg(147)-X-Phe(149)) that were essential for fumarate-dependent transcriptional regulation. The presence of fumarate or d-tartrate caused sharpening of peaks or chemical shift changes in HSQC NMR spectra of the isolated C(4)-dicarboylate binding domain. The amino acid residues responding to fumarate or d-tartrate were in the region comprising residues 89-150 and including the supposed binding site. DcuS(R147A) mutant with an inactivated binding site was isolated and reconstituted in liposomes. The protein showed the same (activation-independent) kinase activity as DcuS, but autophosphorylation of DcuS was no longer stimulated by C(4)-dicarboxylates. Therefore, the R147A mutation affected signal perception and transfer to the kinase but not the kinase activity per se.
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PMID:The nature of the stimulus and of the fumarate binding site of the fumarate sensor DcuS of Escherichia coli. 1578 52

The CheA histidine kinase initiates the signal transduction pathway of bacterial chemotaxis by autophosphorylating a conserved histidine on its phosphotransferase domain (P1). Site-directed mutations of neighboring conserved P1 residues (Glu-67, Lys-48, and His-64) show that a hydrogen-bonding network controls the reactivity of the phospho-accepting His (His-45) in Thermotoga maritima CheA. In particular, the conservative mutation E67Q dramatically reduces phosphotransfer to P1 without significantly affecting the affinity of P1 for the CheA ATP-binding domain. High resolution crystallographic studies revealed that although all mutants disrupt the hydrogen-bonding network to varying degrees, none affect the conformation of His-45. 15N-NMR chemical shift studies instead showed that Glu-67 functions to stabilize the unfavored N(delta1)H tautomer of His-45, thereby rendering the N(epsilon2) imidazole unprotonated and well positioned for accepting the ATP phosphoryl group.
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PMID:Structural and chemical requirements for histidine phosphorylation by the chemotaxis kinase CheA. 1599 28

FixL is a heme-based O(2) sensor, in which the autophosphorylation is regulated by the binding of exogenous ligands such as O(2) and CN(-). In this study, mutants of the heme distal Arg200, Arg208, Ile209, Ile210, and Arg214 residues of SmFixL were characterized biochemically and physicochemically, because it has been suggested that they are significant residues in ligand-linked kinase regulation. Measurements of the autoxidation rate, affinities, and kinetics of ligand binding revealed that all of the above residues are involved in stabilization of the O(2)-heme complex of FixL. However, Arg214 was found to be the only residue that is directly relevant to the ligand-dependent regulation of kinase activity. Although the wild type and R214K and R214Q mutants exhibited normal kinase regulation, R214A, R214M, R214H, and R214Y did not. (13)C and (15)N NMR analyses for (13)C(15)N(-) bound to the truncated heme domains of the Arg214 mutants indicated that, in the wild type and the foregoing two mutants, the heme moiety is present in a single conformation, but in the latter four, the conformations fluctuate possibly because of the lack of an interaction between the iron-bound ligand and residue 214. It is likely that such a rigid conformation of the ligand-bound form is important for the downregulation of histidine kinase activity. Furthermore, a comparison of the NMR data between the wild type and R214K and R214Q mutants suggests that a strong electrostatic interaction between residue 214 and the iron-bound ligand is not necessarily required for the single convergent structure and eventually for the downregulation of FixL.
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PMID:Roles of the heme distal residues of FixL in O2 sensing: a single convergent structure of the heme moiety is relevant to the downregulation of kinase activity. 1648 44

The Rcs signalling pathway controls a variety of physiological functions like capsule synthesis, cell division or motility in prokaryotes. The Rcs regulation cascade, involving a multi-step phosphorelay between the two membrane-bound hybrid sensor kinases RcsC and RcsD and the global regulator RcsB, is, up to now, one of the most complicated regulatory systems in bacteria. To understand the structural basis of Rcs signal transduction, NMR spectroscopy was employed to determine the solution structure of the RcsC C terminus, possessing a phosphoreceiver domain (RcsC-PR), and a region previously described as a long linker between the histidine kinase domain of RcsC (RcsC-HK) and the RcsC-PR. We have found that the linker region comprises an independent structural domain of a new alpha/beta organization, which we named RcsC-ABL domain (Alpha/Beta/Loop). The ABL domain appears to be a conserved and unique structural element of RcsC-like kinases with no significant sequence homology to other proteins. The second domain of the C terminus, the RcsC-PR domain, represents a well-folded CheY-like phosphoreceiver domain with the central parallel beta-sheet covered with two alpha-helical layers on both sides. We have mapped the interaction of RcsC-ABL and RcsC-PR with the histidine phosphotransfer domain (HPt) of RcsD. In addition we have characterized the interaction with and the conformational effects of Mg2+ and the phosphorylation mimetic BeF(-)(3) on RcsC-ABL and RcsC-PR.
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PMID:A new structural domain in the Escherichia coli RcsC hybrid sensor kinase connects histidine kinase and phosphoreceiver domains. 1700 98

The modeling of protein-protein complexes greatly benefits from the incorporation of experimental distance restraints. Pulsed dipolar electron spin resonance spectroscopy is one such powerful technique for obtaining long-range distance restraints in protein complexes. Measurements of the dipolar interaction between two spins placed specifically within a protein complex give information about the spin-spin separation distance. We have developed a convenient method to incorporate such long-range distance information in the modeling of protein-protein complexes that is based on rigid body refinement of the protein components with the software Crystallography and NMR System (CNS). Factors affecting convergence such as number of restraints, error allocation scheme, and number and position of spin labeling sites were investigated with real and simulated data. The use of 4 to 5 different labeling sites on each protein component was found to provide sufficient coverage for producing accuracies limited by the uncertainty in the spin-label conformation within the complex. With an asymmetric scheme of allocating this uncertainty, addition of simulated restraints revealed the importance of longer distances within a limited set of total restraints. We present two case studies: (1) refinement of the complex formed between the histidine kinase CheA and its coupling protein CheW, and (2) refinement of intra-helical separations in the protein a-synuclein bound to micelles.
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PMID:Rigid body refinement of protein complexes with long-range distance restraints from pulsed dipolar ESR. 1760 28

EnvZ is an osmosensing histidine kinase located in the inner membrane, and one of the most extensively studied Escherichia coli histidine kinases. Because of its structural complexity, functional and structural studies have been quite challenging. It is a multidomain transmembrane protein consisting of 450 amino acid residues. In addition, it must form a dimer to function as a histidine kinase like all the other histidine kinases. EnvZ consists of the 115-residue periplasmic domain, two transmembrane domains (TM1 and TM2), and the cytoplasmic domain consisting of the 43-residue linker (HAMP) domain and the 228-residue kinase domain. It has been shown that the kinase domain of EnvZ, responsible for its enzymatic activities, contains all of the conserved regions of histidine kinases such as H, F, N, G1, G2, and G3 boxes. Therefore, the 271-residue cytoplasmic domain of EnvZ (termed EnvZc) has been used as a model system to establish fundamental characteristics of histidine kinases. The DNA fragment encoding EnvZc was cloned in pET vector and EnvZc was expressed and purified. It is highly soluble and retains all the enzymatic activities of EnvZ. We demonstrated that it consists of two functional domains, domain A and domain B. NMR spectroscopic studies of these two domains revealed, for the first time, the structure of a histidine kinase. Domain A is responsible for dimerization of EnvZc forming a four-helical bundle containing two alpha-helical hairpin structures, while domain B is a monomer and has an ATP-binding pocket formed by regions conserved among the histidine kinases. In this chapter, we describe functional and structural studies of EnvZc, which can be applied to characterize other histidine kinases.
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PMID:Functional and structural characterization of EnvZ, an osmosensing histidine kinase of E. coli. 1760 32


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