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)

Chemoreceptor Trg and osmosensor EnvZ of Escherichia coli share a common transmembrane organization but have essentially unrelated primary structures. We created a hybrid gene coding for a protein in which Trg contributed its periplasmic and transmembrane domains as well as a short cytoplasmic segment and EnvZ contributed its cytoplasmic kinase/phosphatase domain. Trz1 transduced recognition of sugar-occupied, ribose-binding protein by its periplasmic domain into activation of its cytoplasmic kinase/phosphatase domain as assessed in vivo by using an ompC-lacZ fusion gene. Functional coupling of sugar-binding protein recognition to kinase/phosphatase activity indicates shared features of intramolecular signalling in the two parent proteins. In combination with previous documentation of transduction of aspartate recognition by an analogous fusion protein created from chemoreceptor Tar and EnvZ, the data indicate a common mechanism of transmembrane signal transduction by chemoreceptors and EnvZ. Signalling through the fusion proteins implies functional interaction between heterologous domains, but the minimal sequence identity among relevant segments of EnvZ, Tar, and Trg indicates that the link does not require extensive, specific interactions among side chains. The few positions of identity in those three sequences cluster in transmembrane segment 1 and the short chemoreceptor sequence in the cytoplasmic part of the hybrid proteins. These regions may be particularly important in physical and functional coupling. The specific cellular conditions necessary to observe ligand-dependent activation of Trz1 can be understood in the context of the importance of phosphatase control in EnvZ signalling and limitations on maximal receptor occupancy in binding protein-mediated recognition.
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PMID:Transmembrane signalling by a hybrid protein: communication from the domain of chemoreceptor Trg that recognizes sugar-binding proteins to the kinase/phosphatase domain of osmosensor EnvZ. 810 26

Protein histidine kinase (Motojima, K., and Goto, S. (1993) FEBS Lett. 319, 75-79) and phosphatase in rat liver extract were characterized. The histidine kinase was recovered mostly in the membrane and the phosphatase in the soluble fraction. The kinase and its substrate 36-kDa protein (P36) were co-solubilized from the membrane under conditions in which most of the other kinases, and their substrate proteins were not solubilized. The solubilized kinase and P36 were co-eluted after high pressure liquid chromatography gel filtration, showing an apparent molecular mass of 70-75 kDa. They were also co-eluted after ion exchange chromatography. These characteristics, together with its complete resistance to genistein, indicate that the rat liver histidine kinase is not cognate to the yeast enzyme (Huang, J., Nasr, M., Kim, Y., and Matthews, H.R. (1992) J. Biol. Chem. 267, 15511-15515). The phosphatase that dephosphorylates histidyl-phosphorylated P36 was also studied using rat liver subcellular fractions and in vitro phosphorylated P36 as the substrate. The characteristics of the phosphatase, that is, 1) Mg2+ requirement for activity, 2) apparent molecular mass of 45 kDa by high performance liquid chromatography gel filtration, and 3) resistance to 100 microM okadaic acid, suggest that the primary phosphatase active in vitro is protein phosphatase 2C.
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PMID:Histidyl phosphorylation and dephosphorylation of P36 in rat liver extract. 813 40

Taz1 is a hybrid receptor in the Escherichia coli cytoplasmic membrane, consisting of the N-terminal ligand binding domain of Tar (a chemoreceptor for aspartate) and the C-terminal signaling domain of EnvZ (an osmosensor). The binding of aspartate to an extra cytoplasmic domain induces the transmembrane signal to the cytoplasmic signaling domain. The signaling domain functioning as a protein kinase evokes a response by transferring a phosphate from an intracellular histidine to OmpR. This domain also encodes an OmpR-specific phosphatase whose action is crucial in completing the OmpR phosphorylation cycle. Phosphorylated OmpR acts as a transcriptional activator for the ompC gene. A number of mutations were introduced into the signaling domain in conserved sequences of the prokaryotic histidine kinase family. All Taz1 mutants lost the ability to both autophosphorylate the histidine residue and transfer the phosphate to OmpR. These mutated receptors were unable to activate ompC-lacZ expression. However, ompC-lacZ was able to be activated by complementation of Taz1 mutants. In some combinations, two different defective Taz1 mutants could restore both OmpR kinase and phosphatase activities when co-expressed. In other combinations only kinase activity was restored. Aspartate-inducible ompC-lacZ expression was restored only in the former cases, while in the latter cases ompC-lacZ expression became constitutive. These results indicate that the kinase activity is essential to activate ompC expression while the phosphatase activity is required to regulate ompC gene expression in a ligand-dependent manner.
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PMID:Requirement of both kinase and phosphatase activities of an Escherichia coli receptor (Taz1) for ligand-dependent signal transduction. 838 84

Taz1 is a hybrid receptor, in which the periplasmic receptor domain of Tar, an aspartate chemoreceptor, is fused with the cytoplasmic signaling domain of EnvZ, an osmosensor. Taz1 is able to induce ompC-lacZ expression in response to aspartate added to the medium. We introduced amino acid substitution mutations in the highly conserved region of the signaling domain of Tar near the Tar-EnvZ junction. The same mutations in Tsr, a serine chemoreceptor, are known to lock the flagella rotation in either a clockwise (CW) or in a counter-clockwise (CCW) mode. It was found that a CW-biased mutation in Taz1 resulted in ompC-lacZ expression in the "off mode", or low ompC-lacZ expression in both the absence and presence of aspartate, while CCW-biased mutations caused ompC-lacZ expression in the "on mode", or constitutive expression regardless of aspartate. The OmpR kinase and phospho-OmpR phosphatase activities of the wild-type and mutant Taz proteins were also examined in response to aspartate. The phosphatase activity of the wild-type Taz1 was found to decrease in the presence of aspartate, while the OmpR kinase activity remained constant. This indicated that aspartate binding to the Taz1 receptor domain modulates the ratio of kinase to phosphatase activity of the signaling domain. An increased kinase to phosphatase ratio in the presence of aspartate resulted in higher levels of phospho-OmpR in the cell and therefore induced ompC-lacZ expression. In contrast to the wild-type Taz1 protein, the enzymatic activities of CW as well as CCW mutants did not change in response to aspartate, indicating that mutant Taz proteins are incapable of transducing the signal across the membrane as a result of a locked conformation of the signaling domain in either the on or off mode.
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PMID:Ligand binding to the receptor domain regulates the ratio of kinase to phosphatase activities of the signaling domain of the hybrid Escherichia coli transmembrane receptor, Taz1. 839 37

Eukaryotic cellular proteins contain phosphohistidine. To search for protein histidine phosphatases, protein histidine kinase from Saccharomyces cerevisiae was used to phosphorylate histone H4 on histidine at position 75 in the H4 amino acid sequence. Incubation of the phosphorylated histone H4 with either protein phosphatase 1, 2A, or 2C resulted in extensive removal of phosphate from the phosphorylated histone. Thus, protein phosphatases 1, 2A, and 2C are histidine phosphatases as well as serine/threonine phosphatases. Calcium/calmodulin-regulated protein phosphatase (protein phosphatase 2B) did not remove phosphate from phosphohistidine. The histidine phosphatase reaction was tested for a magnesium requirement and effects of inhibitor-1 and okadaic acid. In all cases, the protein phosphatases behaved as they do in their serine/threonine phosphatase activity. Extracts of the yeast, S. cerevisiae, contain protein histidine phosphatase activity. Quantitative measurement of phosphatase activity shows that the activity against phosphohistidine is a major activity of protein phosphatases 1, 2A, and 2C.
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PMID:Protein phosphatases 1, 2A, and 2C are protein histidine phosphatases. 839 6

EnvZ of Escherichia coli is a transmembrane histidine kinase belonging to the family of two-component signal transducing systems prevalent in prokaryotes and recently discovered in eukaryotes. In response to changes in medium osmolarity EnvZ regulates the level of phosphorylated OmpR, its conjugate response-regulating transcription factor for ompF and ompC genes. EnvZ has dual opposing enzymatic activities; OmpR-phosphorylase (kinase) and phospho-OmpR-dephosphorylase (phosphatase). The osmotic signal is proposed to regulate the ratio of the kinase to the phosphatase activities of EnvZ to modulate the level of OmpR phosphorylation. In this work we used a COOH-terminal fragment of a previously identified kinase-/phosphatase+ EnvZ mutant (EnvZ-N347D) to demonstrate that the phosphoryl group on phospho-OmpR is transferred back to EnvZ to the same histidine residue (His243) that is utilized for the autokinase reaction by the wild type protein. Phospho-EnvZ-N347D thus formed could also transfer its phosphoryl group back to OmpR. The phosphotransfer reaction from phospho-OmpR to EnvZ.N347D was inhibited by ADP while Mg2+ ions stimulated the dephosphorylation reaction, resulting in release of inorganic phosphate. These results indicate that the energy levels of phosphoryl groups on OmpR and EnvZ are very similar and that the phosphatase reaction in the EnvZ.N347D mutant involves a reversal of the phosphotransfer reaction from EnvZ to OmpR using the identical His243 residue.
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PMID:Reverse phosphotransfer from OmpR to EnvZ in a kinase-/phosphatase+ mutant of EnvZ (EnvZ.N347D), a bifunctional signal transducer of Escherichia coli. 857 33

EnvZ undergoes autophosphorylation at His243 and subsequently transfers the phosphate group to OmpR. EnvZ also possesses an OmpR-phosphate phosphatase activity. We examined the role of His243 in the phosphatase function by replacing His with either Val, Tyr, Ser, Asp, or Asn. EnvZH243V and EnvZH243Y were both shown to possess phosphatase activity in vitro. In addition, the mutant proteins were able to reduce the high level of OmpR-phosphate present in the envZ473 strain. These results indicate that His243 of EnvZ is not essential for stimulating the dephosphorylation of OmpR-phosphate.
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PMID:Role of His243 in the phosphatase activity of EnvZ in Escherichia coli. 902 31

EnvZ and OmpR are the sensor and response regulator proteins of a two-component system that controls the porin regulon of Escherichia coli in response to osmolarity. Three enzymatic activities are associated with EnvZ: autokinase, OmpR kinase, and OmpR-phosphate (OmpR-P) phosphatase. Conserved histidine-243 is critical for both autokinase and OmpR kinase activities. To investigate its involvement in OmpR-P phosphatase activity, histidine-243 was mutated to several other amino acids and the phosphatase activity of mutated EnvZ was measured both in vivo and in vitro. In agreement with previous reports, we found that certain substitutions abolished the phosphatase activity of EnvZ. However, a significant level of phosphatase activity remained when histidine-243 was replaced with certain amino acids, such as tyrosine. In addition, the phosphatase activity of a previously identified kinase- phosphatase+ mutant was not abolished by the replacement of histidine-243 with asparagine. These data indicated that although conserved histidine-243 is important for the phosphatase activity, a histidine-243-P intermediate is not required. Our data are consistent with a previous model that proposes a common transition state with histidine-243 (EnvZ) in close contact with aspartate-55 (OmpR) for both OmpR phosphorylation and dephosphorylation. Phosphotransfer occurs from histidine-243-P to aspartate-55 during phosphorylation, but water replaces the phosphorylated histidine side chain leading to hydrolysis during dephosphorylation.
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PMID:Function of conserved histidine-243 in phosphatase activity of EnvZ, the sensor for porin osmoregulation in Escherichia coli. 917 23

EnvZ, a transmembrane signal transducer, is composed of a periplasmic sensor domain, transmembrane domains, and a cytoplasmic signaling domain. Between the second transmembrane domain and the cytoplasmic signaling domain there is a linker domain consisting of approximately 50 residues. In this study, we investigated the functional role of the EnvZ linker domain with respect to signal transduction. Amino acid sequence alignment of linker regions among various bacterial signal transducer proteins does not show a high sequence identity but suggests a common helix 1-loop-helix 2 structure. Among several mutations introduced in the EnvZ linker region, it was found that hydrophobic-to-charged amino acid substitutions in helix 1 and helix 2 and deletions in helix 1, loop, and helix 2 (delta14, delta8, and delta7) resulted in constitutive OmpC expression. In the linker mutant EnvZ x delta7, both kinase and phosphatase activities were significantly reduced but the ratio of kinase to phosphatase activity increased, consistent with the constitutive OmpC expression. In contrast, the purified cytoplasmic fragment of EnvZ x delta7 possessed both kinase and phosphatase activities at levels similar to those of the cytoplasmic fragment of wild-type EnvZ. In addition, the linker mutations had no direct effect on EnvZ C-terminal dimerization. These results together with previous data suggest that the linker region is not directly involved in EnvZ enzymatic activities and that it may have a crucial role in propagating a conformational change to ensure correct positioning of two EnvZ molecules within a dimer during the transmembrane signaling.
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PMID:Mutational analysis of the linker region of EnvZ, an osmosensor in Escherichia coli. 920 57

Disruption of normal protein trafficking in the Escherichia coli cell envelope (inner membrane, periplasm, outer membrane) can activate two parallel, but distinct, signal transduction pathways. This activation stimulates the expression of a number of genes whose products function to fold or degrade the mislocalized proteins. One of these signal transduction pathways is a two-component regulatory system comprised of the histidine kinase CpxA and the response regulator, CpxR. In this study we characterized gain-of-function Cpx* mutants in order to learn more about Cpx signal transduction. Sequencing demonstrated that the cpx* mutations cluster in either the periplasmic, the transmembrane, or the H-box domain of CpxA. Intriguingly, most of the periplasmic cpx* gain-of-function mutations cluster in the central region of this domain, and one encodes a deletion of 32 amino acids. Strains harboring these mutations are rendered insensitive to a normally activating signal. In vivo and in vitro characterization of maltose-binding-protein fusions between the wild-type CpxA and a representative cpx* mutant, CpxA101, showed that the mutant CpxA is altered in phosphotransfer reactions with CpxR. Specifically, while both CpxA and CpxA101 function as autokinases and CpxR kinases, CpxA101 is devoid of a CpxR-P phosphatase activity normally present in the wild-type protein. Taken together, the data support a model for Cpx-mediated signal transduction in which the kinase/phosphatase ratio is elevated by stress. Further, the sequence and phenotypes of periplasmic cpx* mutations suggest that interactions with a periplasmic signaling molecule may normally dictate a decreased kinase/phosphatase ratio under nonstress conditions.
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PMID:Transduction of envelope stress in Escherichia coli by the Cpx two-component system. 940 Oct 31


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