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)

OmpR is a DNA-binding protein that regulates transcription of ompF and ompC. The activity of OmpR is controlled by the inner membrane osmosensor, EnvZ. In order to study the signaling process between EnvZ and OmpR, we analyzed two different envZ strains: the envZ473 strain, in which OmpC is constitutively produced and OmpF is fully repressed, and the envZ3 strain, in which the production of OmpC is greatly reduced and OmpF is not fully repressed by high-osmolarity growth conditions. Using direct sequencing of DNA derived from the polymerase chain reaction amplification method, we identified the mutation in the envZ473 strain as a Val-241-to-Gly substitution and the mutation in the envZ3 as an Ala-219-to-Val substitution. The relative DNA-binding affinity of OmpR derived from the envZ473 strain was dramatically increased for the upstream sequence of both ompF and ompC. In contrast, OmpR derived from the envZ3 strain was not converted to the high-affinity form. The intracellular levels of OmpR-phosphate, as analyzed by the in vivo phosphorylation approach, significantly increased in the envZ473 strain, while in the envZ3 strain the levels were considerably reduced, relative to those found in the parent strain. The intracellular level of OmpR protein in the envZ473 strain was also found to be markedly elevated relative to that of the parent strain. These results are discussed in relation to the role of phosphorylation and relative DNA-binding affinity of OmpR in the expression of ompF and ompC.
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PMID:Molecular analysis of the signaling pathway between EnvZ and OmpR in Escherichia coli. 131 Dec 95

The rates of synthesis of a class of both secreted and intracellular degradative enzymes in Bacillus subtilis are controlled by a signal transduction pathway defined by at least four regulatory genes: degS, degU, degQ (formerly sacQ), and degR (formerly prtR). The DegS-DegU proteins show amino acid similarities with two-component procaryotic modulator-effector pairs such as NtrB-NtrC, CheA-CheY, and EnvZ-OmpR. By analogy with these systems, it is possible that DegS is a protein kinase which could catalyze the transfer of a phosphoryl moiety to DegU, which acts as a positive regulator. DegR and DegQ correspond to polypeptides of 60 and 46 amino acids, respectively, which also activate the synthesis of degradative enzymes. We show that the degS and degU genes are organized in an operon. The putative sigma A promoter of the operon was mapped upstream from degS. Mutations in degS and degU were characterized at the molecular level, and their effects on transformability and cell motility were studied. The expression of degQ was shown to be subject both to catabolite repression and DegS-DegU-mediated control, allowing an increase in the rate of synthesis of degQ under conditions of nitrogen starvation. These results are consistent with the hypothesis that this control system responds to an environmental signal such as limitations of nitrogen, carbon, or phosphate sources.
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PMID:Signal transduction pathway controlling synthesis of a class of degradative enzymes in Bacillus subtilis: expression of the regulatory genes and analysis of mutations in degS and degU. 168 43

Bacterial motility and gene expression are controlled by a family of phosphorylated response regulators whose activities are modulated by an associated family of protein-histidine kinases. In chemotaxis there are two response regulators, CheY and CheB, that receive phosphoryl groups from the histidine kinase, CheA. Here we show that the response regulators catalyze their own phosphorylation in that both CheY and CheB can be phosphorylated in the complete absence of any auxiliary protein. Both CheY and CheB use the N-phosphoryl group in phosphoramidate (NH2PO3(2-)) as a phospho-donor. This enzymatic activity probably reflects the general ability of response regulators to accept phosphoryl groups from phosphohistidines in their associated kinases. It provides a general method for the study of activated response regulators in the absence of kinase proteins. CheY can also use intermediary metabolites such as acetyl phosphate and carbamoyl phosphate as phospho-donors. These reactions may provide a mechanism to modulate cell behavior in response to altered metabolic states.
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PMID:Phosphorylation of bacterial response regulator proteins by low molecular weight phospho-donors. 173 45

The Tar-EnvZ hybrid molecule (Taz1) is an inner membrane transducer that activates OmpR, a transcriptional activator for porin gene expression (ompC), in response to an aspartic acid signal. Signal transduction by Taz1 most likely involves a phosphorylated Taz1 intermediate that donates its phosphate to OmpR. Phosphorylated OmpR has already been implicated in transcriptional activation of porin genes. Using a cell-free system containing Taz1-enriched membrane fractions, we have examined the phosphorylation properties of Taz1 and the stimulatory effects of divalent and monovalent ions. Highest activation of Taz1 phosphorylation was observed with CaCl2, and its stimulation could be observed with as low as 60 microM of CaCl2. Phosphorylated Taz1 could readily donate its phosphate group to OmpR in the presence of calcium. CaCl2 was also able to enhance phosphorylation of intact membrane-bound EnvZ and a cytoplasmic fragment of EnvZ lacking the receptor and transmembrane domains. These results indicate that the site for CaCl2 stimulation is within the cytoplasmic region of EnvZ and probably involves an enhanced rate of EnvZ phosphorylation.
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PMID:Ca2(+)-enhanced phosphorylation of a chimeric protein kinase involved with bacterial signal transduction. 185 Apr 14

An enzyme of molecular weight 32,000 comprising a single subunit has been isolated from whole cell extracts of the yeast Saccharomyces cerevisiae. In vitro, the enzyme transfers the gamma phosphate of ATP to a protein substrate, histone H4, to produce an alkali-stable phosphorylation. Modification of the substrate histidine with diethylpyrocarbonate prevented phosphorylation. Phosphoamino acid analysis of the phosphorylated substrate showed the presence of 1-phosphohistidine. Hence, the isolated enzyme is a protein histidine kinase. A novel assay for acid-labile alkali-stable protein phosphorylation was used in the purification of the kinase activity to a final specific activity of 2,700 nmol/15 min/mg. The purified enzyme phosphorylates specifically histidine 75 in histone H4 and does not phosphorylate histidine 18 nor histidine residues in any other core histone. Steady state kinetic data are consistent with an ordered sequential reaction with Km values for Mg-ATP and histone H4 of 60 and 17 microM, respectively. The protein histidine kinase requires a divalent cation such as Mg2+, Co2+, or Mn2+ but will not use Ca2+, Zn2+, Cu2+, Fe2+, spermine, or spermidine. This is the first purification of an enzyme that catalyzes N-linked phosphorylation in proteins.
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PMID:Purification of a protein histidine kinase from the yeast Saccharomyces cerevisiae. The first member of this class of protein kinases. 202 10

In Escherichia coli, adaptation to hyperosmotic conditions alters the expression of the outer membrane porins OmpF and OmpC. The amount of PhoE porin, which is normally induced by phosphate deprivation, was greatly reduced in cells adapted to high-osmolarity conditions. Osmoregulation of PhoE operated independently of the activity of the PhoR phosphate sensor and did not involve cross-talk from the homologous osmosensor EnvZ. PhoE synthesis was partially restored by additional copies of the positive regulator phoB+ and by the osmoprotectant glycine betaine.
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PMID:Osmotic regulation of PhoE porin synthesis in Escherichia coli. 216 86

Agrobacterium tumefaciens virulence genes are induced by plant signals through the VirA-VirG two-component regulatory system. The VirA protein is a membrane-spanning sensor molecule that possesses an autophosphorylating activity, and the VirG protein is a sequence-specific DNA-binding protein. In this report, we demonstrate that the VirG protein is phosphorylated by the VirA protein and that the phosphate is directly transferred from the phosphorylated VirA molecule (phosphohistidine) to the VirG protein. The chemical stability of the phospho-VirG bond suggested that the VirG protein was phosphorylated at the aspartate and/or glutamate residue. The phosphorylated VirG protein was reduced with tritiated sodium borohydride and subjected to proteolytic digestion with the Achromobacter protease I enzyme. The resulting peptide fragments were separated by C8 reversed-phase high-pressure liquid chromatography, and the tritium-labeled peptide was sequenced. Amino acid sequence data showed that the aspartate residue at position 52 was the only site phosphorylated. Changing this aspartate into asparagine resulted in a nonphosphorylatable and biologically nonfunctional gene product. As a control, a randomly chosen aspartate was changed into an asparagine (position 72), and no effect on its phosphorylation or biological activity was observed. Unlike its homologs, including CheA-CheY, EnvZ-OmpR, and NtrB-NtrC, the phospho-VirG molecule was very stable in vitro. The possible implications of these observations and the function of VirG phosphorylation in vir gene activation are discussed.
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PMID:Phosphorylation of the VirG protein of Agrobacterium tumefaciens by the autophosphorylated VirA protein: essential role in biological activity of VirG. 239 78

EnvZ and OmpR, the regulatory proteins for ompF and ompC expression in Escherichia coli, belong to a modulator-effector family of regulatory proteins which are essential for the response to environmental signals. We show that the soluble cytoplasmic domain of the transmembrane modulator protein EnvZ is phosphorylated in vitro by [gamma-32P]-ATP. We also demonstrate that the phosphate group can, in turn, be transferred to the transcription activator protein OmpR. The pH stability properties of the phosphate groups linked to EnvZ indicate that this molecule contains histidyl phosphate. The invariant His-243 of EnvZ corresponds to the phosphorylated His-48 of the chemotactic modulator protein CheA. Substitution of His-243 with valine produces an EnvZ that is refractory to phosphorylation and can no longer catalyze the transfer of phosphate to OmpR. Furthermore, in a delta envZ strain of E. coli, containing the envZ Val-243 plasmid, ompC expression is elevated 7-fold relative to that found in cells carrying the wild-type envZ plasmid. Based on these results we propose a model in which the phosphorylated state of OmpR modulates the expression of the ompF and ompC genes.
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PMID:Phosphorylation of OmpR by the osmosensor EnvZ modulates expression of the ompF and ompC genes in Escherichia coli. 266 53

The phoB gene product of Escherichia coli is the transcriptional activator for the genes in the phosphate regulon as well as for phoB itself, all of which are induced by phosphate starvation. The phoR gene product modulates PhoB function in response to the phosphate concentrations in the medium. We quantitatively compared the levels of expression of the phoA, phoB, phoE, and pstS genes in several phoB mutants with different phenotypes by constructing operon fusions of these genes with the gene for chloramphenicol acetyltransferase. Although all the phoB mutants examined had little activator function for phoA, three among the four mutants showed various levels of the activator function for phoB, pstS, and phoE. To study the functional motifs of the PhoB and PhoR proteins, we cloned and sequenced the four classical phoB and six phoR mutant genes. All of the phoB mutations and one of the phoR mutations were missense mutations, and most of the altered amino acids were in the highly conserved amino acids among the regulatory proteins homologous to PhoB or PhoR protein, such as the OmpR, SfrA, and VirG proteins or the EnvZ, CpxA, and VirA proteins. The other five phoR mutations were nonsense mutations.
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PMID:Regulation of the phosphate regulon of Escherichia coli: analysis of mutant phoB and phoR genes causing different phenotypes. 267 81

Hexokinase (EC 2.7.1.1) catalyzes the first step in glucose metabolism, using ATP for the phosphorylation of glucose to glucose 6-phosphate. A portion of the HK1 gene was cloned by mixed oligonucleotide primer amplification of cDNA using primers of high complexity. The amino acid sequence for a partial fragment of bovine cardiac muscle HK was determined and used to create primer mixtures of 256- and 1024-fold complexity. Two products were generated from bovine cardiac muscle cDNA which show 82% nucleotide and 93% amino acid identity with a region of rat brain HK1 and cDNA. This work demonstrates that extension and amplification of cDNA probes may be successful even when amino acid sequence data indicate substantial codon degeneracy.
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PMID:Synthesis and characterization of a bovine hexokinase 1 cDNA probe by mixed oligonucleotide primed amplification of cDNA using high complexity primer mixtures. 271 57


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