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)

K+ channels represent the most complex class of voltage-gated ion channels from both functional and structural standpoints. In the heart these channels are responsible for the rapid repolarizing phases of the action potential and are the targets of several antiarrhythmic drugs. Full-length cDNA clones were isolated from human ventricular libraries that encode two voltage-gated K+ channels. These two cDNAs, designated HK1 and HK2, encode proteins of 653 and 605 amino acids, respectively. HK1 is the human equivalent (98% identity) of an inactivating K+ channel previously described in rat heart (RHK1) whereas the HK2 channel is 86% identical to a cloned rat brain K+ channel (Kv1). The only amino acid sequence identity (72%) between HK1 and HK2 is within the central region containing the membrane spanning domains. Northern blot analysis of human mRNA indicated that HK1 is slightly more abundant in ventricle than atrium whereas HK2 is much more abundant in atrium relative to ventricle. Both channel transcripts are present in ventricle at levels equivalent to voltage-gated Na+ channels. Analysis of the gene encoding HK1 suggests the coding sequence is intronless and is represented once in the human genome.
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PMID:Molecular cloning and characterization of two voltage-gated K+ channel cDNAs from human ventricle. 200 94

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

The transcriptional factors, OmpR and EnvZ, are crucially involved in the osmotic regulation of ompF and ompC expression in Escherichia coli. The DNA binding ability of the positive regulator, OmpR, is modulated through its phosphorylation and dephosphorylation mediated by EnvZ in response to the medium osmolarity. In this study, two examples of a novel type of mutant ompR allele, ompR96A and ompR115S, whose phenotype is OmpF- OmpC- irrespective of the medium osmolarity, were characterized. These mutations result in amino acid conversions, Glu96 to Ala and Arg115 to Ser, respectively, within the phosphorylation domain of OmpR. Nevertheless, these mutant proteins were capable of undergoing phosphorylation and dephosphorylation normally, just like wild-type OmpR. However, the phosphorylation-dependent enhancement of their in vitro DNA binding ability was found to be severely affected. It was thus revealed that these mutant OmpR represent a novel type in terms of the mechanism of phosphorylation-dependent activation of the function of OmpR, i.e. those are normally phosphorylated but not activated to bind to the cognate promoter DNAs. In this respect, it was further suggested that OmpR oligomerization may be involved in the mechanism underlying the phosphorylation-dependent enhancement of the DNA binding ability of OmpR. The mutant proteins characterized in this study seem to be defective in this particular oligomerization process observed in vitro.
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PMID:Signal transduction and osmoregulation in Escherichia coli. A novel type of mutation in the phosphorylation domain of the activator protein, OmpR, results in a defect in its phosphorylation-dependent DNA binding. 204 May 97

An in vivo approach was taken to assess whether the phosphorylated state of the transcription activator OmpR was affected by changes in the osmolarity of the growth medium or by mutations in envZ, the gene encoding the inner membrane histidine kinase that phosphorylates OmpR. We present results that support the view that increased phosphorylation of OmpR is correlated with enhanced expression of ompC. The in vivo phosphorylation approach was also used to show that OmpR can be phosphorylated in an envZ null strain. This result indicates that phosphorylation cross talk can occur in vivo between OmpR and a kinase(s) that is functionally homologous to envZ.
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PMID:In vivo phosphorylation of OmpR, the transcription activator of the ompF and ompC genes in Escherichia coli. 216 Sep 45

Ly-6C is a differentiation antigen that distinguishes T-lymphocyte subsets. In concordance with previous results, splenocytes from NOD mice do not express the epitope recognized by anti-Ly-6C monoclonal antibodies (MoAbs), including MoAb HK1.4 in this study, and cannot be stimulated to proliferate in response to HK1.4. However, when splenocytes from NOD mice were stimulated in vitro with the anti-CD3 MoAb 145-2C11, T lymphocytes expressing Ly-6C were detected after 48 h of stimulation, with as many as 25% of lymphocytes expressing this antigen with prolonged passage in culture. Most of the cells expressing Ly-6C were Thy-1.2+, CD4+, and CD8- and proliferated after stimulation with HK1.4. To further understand the failure of NOD splenocytes to express Ly-6C, freshly isolated cells were stimulated with alpha/beta-interferon (IFN-alpha/beta) and IFN-gamma. Although these lymphokines induced expression of Ly-6A and Ly-6C in splenocytes from C57BL/6J mice and Ly-6A in NOD cells, Ly-6C was not induced on NOD cells. Because Ly-6C expression on splenocytes was a marker of activation via the CD3 T-lymphocyte receptor complex, we also examined expression of Ly-6C on T lymphocytes within islets showing insulitis in vivo. Lymphocytes that were Ly-6C+ were identified within islets on histological sections of pancreas, whereas Ly-6C+ cells in the spleen from the same mouse could not be detected. Our findings imply functional abnormality in expression of Ly-6C in NOD mice.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Expression of Ly-6C by T lymphocytes of NOD mice after CD3-complex stimulation. Identification of activated cells during insulitis of prediabetic mice. 216 2

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

Previously, the transfer of a phosphoryl group between the EnvZ and OmpR proteins, which are involved in expression of the ompF and ompC genes in response to the medium osmolarity, was demonstrated in vitro. In this study, the histidine (His) residue at position 243 of the EnvZ protein, and the aspartate (Asp) residues at positions 12 and 55 of the OmpR protein were changed, respectively, by means of site-directed mutagenesis. We characterized the mutant proteins in terms of not only their in vitro phosphotransfer reactions but also their in vivo osmoregulatory phenotypes. The mutant EnvZ protein was defective in its in vitro ability not only as to EnvZ-autophosphorylation but also OmpR-phosphorylation and OmpR-dephosphorylation. This particular mutant EnvZ protein seemed to exhibit null functions as to the in vivo osmoregulatory phenotype. The mutant OmpR protein with the amino acid change at position 12 was clearly phosphorylated in vitro, but at a very low rate as compared with the wild-type OmpR protein. In vitro phosphorylation of the mutant OmpR protein with the amino acid change at position 55 was more severely affected. This mutant OmpR protein appeared to exhibit null functions as to the in vivo osmoregulatory phenotype. These results suggest that the histidine residue at position 243 of the EnvZ protein and the aspartate residues at positions 12 and 55 of the OmpR protein are deeply involved in the phosphotransfer between the EnvZ and OmpR proteins.
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PMID:Transmembrane signal transduction and osmoregulation in Escherichia coli: I. Analysis by site-directed mutagenesis of the amino acid residues involved in phosphotransfer between the two regulatory components, EnvZ and OmpR. 227 41

The EnvZ protein is presumably a membrane-located osmotic sensor, which specifically phosphorylates the activator protein, OmpR, involved in expression of the ompF and ompC genes in Escherichia coli. In this study, we developed an in vitro system for analyzing the intact form of the EnvZ protein located in the isolated cytoplasmic membrane. This particular form of the EnvZ protein exhibited its in vitro ability not only as to OmpR-phosphorylation but also OmpR-dephosphorylation. It was found that when a high concentration of a mono-cation (K+, Na-, or Li+) was present during the in vitro reactions, OmpR-dephosphorylation was preferentially inhibited and consequently the phosphorylated from of the OmpR protein was accumulated under the in vitro conditions used, although the K+ ion appears to be essential for the OmpR-phosphorylation reaction. Procaine, a local anesthetic, is known to affect the osmotic regulation of the ompF and ompC genes in vivo. In this study, procaine was also found to preferentially inhibit OmpR-dephosphorylation mediated by the EnvZ protein in vitro.
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PMID:Transmembrane signal transduction and osmoregulation in Escherichia coli: II. The osmotic sensor, EnvZ, located in the isolated cytoplasmic membrane displays its phosphorylation and dephosphorylation abilities as to the activator protein, OmpR. 227 42

Alkali-stable phosphorylation of proteins, particularly phosphotyrosine and phosphohistidine, is an important phenomenon in cells. In the case of phosphohistidine and some other phosphoamino acids, the phosphorylation is acid-labile and in these cases studies have been severely limited by the absence of a rapid assay suitable for acid-labile phosphorylation. The assay presented here involves a conventional kinase assay reaction followed by mild alkaline hydrolysis and adsorption of the product to washed Nytran paper at high pH. After further washing, at pH 9, the radioactivity on the papers is determined by liquid scintillation counting. Hence, acid-labile phosphorylation is preserved. The assay is selective for alkali-stable phosphorylation but not fully specific, mainly due to the need to balance the severity of the partial alkaline hydrolysis with the stability of the protein-peptide bonds. The assay has been used for the purification and characterization of a protein histidine kinase from Saccharomyces cerevisiae.
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PMID:A filter-based protein kinase assay selective for alkali-stable protein phosphorylation and suitable for acid-labile protein phosphorylation. 229 65

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


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