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)

Using insulin-secreting cells, we previously demonstrated that specific proteins associated with the cytosolic, secretory granule, and mitochondrial fractions undergo a novel type of phosphorylation on their histidine residues. Subsequently, we identified these proteins as the nucleoside diphosphate kinase (NDPK) [Kowluru and Metz, Biochemistry 1994;33:12495-503], the beta subunit of trimeric GTP-binding proteins [Kowluru et al., Biochem J 1996;313:97-107], and the alpha subunit of succinyl-CoA synthetase [Kowluru, Diabetologia 2001;44:89-94], respectively. Since several other enzymes of intermediary metabolism (e.g. ATP-citrate lyase and glucose-6-phosphatase) also undergo histidine phosphorylation, these initial findings may have a more generalized significance to beta cells. Herein, we characterized a novel protein histidine kinase in pancreatic beta cells, and determined it to be acid- and heat-labile as well as alkali-resistant in its phosphorylation of histone 4. Such an activity was detected in normal rat islets, human islets, and clonal beta (HIT-T15 and INS-1) cells, and could utilize either ATP or GTP as a phosphoryl donor (with K(m) values in the range of 60-100 microM). On a size-exclusion column, its molecular mass was estimated to be in the range of 60-70 kDa. It was stimulated by divalent cations (Mg(2+)>Mn(2+)>control=Ca(2+)=Zn(2+)=Co(2+)), but was resistant to polyamines. It was inactivated by known in vitro inhibitors of protein histidine phosphorylation (e.g. UDP or cromoglycate). Mastoparan, a global activator of G-proteins and insulin secretion from isolated beta cells, but not mastoparan-17, its inactive analog, stimulated histidine kinase activity and histidine phosphorylation of G(beta) subunit and insulin secretion from isolated rat islets. These studies identify, for the first time, a protein kinase activity in the pancreatic beta cell that does not act on traditional -Ser, -Tyr, or -Thr residues. They also establish a possible link between histidine kinase activity and G(beta) phosphorylation in isolated beta cells.
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PMID:Identification and characterization of a novel protein histidine kinase in the islet beta cell: evidence for its regulation by mastoparan, an activator of G-proteins and insulin secretion. 1211 Mar 68

The 90-kDa heat shock family (HSP90) of protein and two-component histidine kinases, although quite distinct at the primary amino acid sequence level, share a common structural ATP-binding domain known as the Bergerat fold. The Bergerat fold is important for the ATPase activity and associated chaperone function of HSP90. Two-component histidine kinases occur in bacteria, yeast, and plants but have yet to be identified in mammalian cells. The antifungal antibiotic radicicol (Monorden) has been shown to bind to the Bergerat fold of HSP90 and to inhibit its ATPase activity. The structural similarity between the Bergerat fold of HSP90 and bacterial two-component histidine kinases prompted our inquiry into whether radicicol could be a potential inhibitor of histidine kinase-like proteins. Structural homology searches suggest that the ATP-binding domains of the yeast histidine kinase Sln1 and the mammalian, branched-chain alpha-keto acid dehydrogenase kinase are very similar to that of other Bergerat fold family members. On the basis of structural homology, we tested radicicol as a potential inhibitor of Sln1 and branched-chain alpha-keto acid dehydrogenase kinase (BCKDHK) and propose a mechanism of inhibition of these kinases. Although BCKDHK has been shown to have serine autophosphorylation activity, we speculate, based on the results from this study and other supporting evidence, that BCKDHK may also have intrinsic histidine kinase activity.
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PMID:Inhibition of branched-chain alpha-keto acid dehydrogenase kinase and Sln1 yeast histidine kinase by the antifungal antibiotic radicicol. 1213 Jun 80

Histidine kinase EnvZ, a transmembrane osmotic sensor for Escherichia coli, is a bifunctional enzyme having OmpR (its cognate response regulator) kinase and phosphorylated OmpR (OmpR-P) phosphatase activities. Its cytoplasmic domain consists of domain A responsible for dimerization of EnvZ, histidine phosphotransfer and phosphatase activities, and domain B responsible for ATP binding. Here, we have constructed a number of substitution mutations at the G2 box, one of the conserved motifs in domain B, and demonstrated that they influence the phosphatase activity of EnvZ over a wide range. The effects of ADP, a cofactor for the phosphatase activity, were found to be substantially different depending upon the mutations. The effects of these mutations were also examined in vivo using a chimeric Tar-EnvZ construct (Taz1-1), and the results agreed with the in vitro data for the phosphatase and kinase activities for all mutations. Using Taz1-1 carrying the T402A mutation, three independent intragenic suppressor mutations (T235M, S269L and E276K) were isolated, and all were found in domain A. Together, the present results demonstrate for the first time that domain A and domain B are functionally co-ordinated and topologically arranged in a specific manner. The G2 box may modulate the interaction between these two domains in response to extracellular osmolarity.
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PMID:The role of the G2 box, a conserved motif in the histidine kinase superfamily, in modulating the function of EnvZ. 1213 13

Two-component and phosphorelay signal transduction systems are the major means by which bacteria recognize and respond to a variety of environmental stimuli. Although several model systems, including sporulation in Bacillus subtilis and chemotaxis in Escherichia coli, have been extensively studied, the two-component signal transduction systems in industrially important actinomycetes are not well studied. We report the molecular and biochemical characterization of a novel two-component signal system, amrA-amkA,from the rifamycin-SV-producing Amycolatopsis mediterranei U32. The deduced sequences of amkAand amrA contain all the structural features that are highly conserved in the typical bacterial histidine kinases and response regulators, respectively. BLAST analyses showed that AmrA and AmkA displayed high similarities to AfsQ1/AfsQ2 of Streptomyces coelicolor and MtrA/MtrB of Mycobacterium tuberculosis. The amrAand amkA genes were over-expressed and the gene products were purified from E. coli. Biochemical studies showed that AmkA is able to autophosphorylate, supporting its functional assignment as a histidine kinase. That AmrA functions as the cognate response regulator for histidine kinase AmkA was demonstrated by in vitro phosphotransfer from [gamma-(32)P]ATP-labeled AmkA to AmrA. Rifamycin SV production was also decreased by 10-20% in amrAor amkA gene disruption mutants under the tested condition. Although the detailed regulatory mechanism is still unknown, this is the first report regarding the involvement of two-component signal systems in rifamycin biosynthesis in the genus Amycolatopsis.
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PMID:Molecular and biochemical characterization of a novel two-component signal transduction system, amrA- amkA, involved in rifamycin SV production in Amycolatopsis mediterranei U32. 1237 6

EnvZ is a histidine protein kinase important for osmoregulation in bacteria. While structural data are available for this enzyme, the nucleotide binding pocket is not well characterized. The ATP binding domain (EnvZB) was expressed, and its ability to bind nucleotide derivatives was assessed using equilbrium and stopped-flow fluorescence spectroscopy. The fluorescence emission of the trinitrophenyl derivatives, TNP-ATP and TNP-ADP, increase upon binding to EnvZB. The fluorescence enhancements were quantitatively abolished in the presence of excess ADP, indicating that the fluorescent probes occupy the nucleotide binding pocket. Both TNP-ATP and TNP-ADP bind to EnvZB with high affinity (K(d) = 2-3 microM). The TNP moiety attached to the ribose ring does not impede access of the fluorescent nucleotide into the binding pocket. The association rate constant for TNP-ADP is 7 microM(-1) s(-1), a value consistent with those for natural nucleotides and the eucaryotic protein kinases. Using competition experiments, it was found that ATP and ADP bind 30- and 150-fold more poorly, respectively, than the corresponding TNP-derivatized forms. Surprisingly, the physiological metal Mg(2+) is not required for ADP binding and only enhances ATP affinity by 3-fold. Although portions of the nucleotide pocket are disordered, the recombinant enzyme is highly stable, unfolding only at temperatures in excess of 70 degrees C. The unusually high affinity of the TNP derivatives compared to the natural nucleotides suggests that hydrophobic substitutions on the ribose ring enforce an altered binding mode that may be exploited for drug design strategies.
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PMID:Probing the nucleotide binding domain of the osmoregulator EnvZ using fluorescent nucleotide derivatives. 1243 44

A histidine kinase-based signaling system has been proposed to function in ethylene signal transduction pathway of plants and one ethylene receptor has been found to possess His kinase activity. Here we demonstrate that a His kinase-like ethylene receptor homologue NTHK1 from tobacco has serine/threonine (Ser/Thr) kinase activity, but no His kinase activity. Evidence obtained by analyzing acid/base stability, phosphoamino acid and substrate specificity of the phosphorylated kinase domain, supports this conclusion. In addition, mutation of the presumptive phosphorylation site His (H378) to Gln did not affect the kinase activity whereas deletion of the ATP-binding domain eliminated it, indicating that the conserved His (H378) is not required for the kinase activity and this activity is intrinsic to the NTHK1-KD. Moreover, confocal analysis of NTHK1 expression in insect cells and plant cells suggested the plasma membrane localization of the NTHK1 protein. Thus, NTHK1 may represent a distinct Ser/Thr kinase-type ethylene receptor and function in an alternative mechanism for ethylene signal transduction.
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PMID:Serine/threonine kinase activity in the putative histidine kinase-like ethylene receptor NTHK1 from tobacco. 1253 51

Some cyanobacteria couple oxygenic photosynthesis in vegetative cells with O2-sensitive N2 fixation in differentiated cells called heterocysts. Heterocyst differentiation involves extensive biochemical and structural changes that collectively permit heterocysts to assimilate N2 aerobically and supply the products of N2 fixation to vegetative cells. HepK and DevR are required for the development of functional heterocysts in Anabaena and Nostoc, respectively. We show that HepK is an autokinase and that Anabaena DevRA is its cognate response regulator, together comprising part or all of a two-component system that mediates developmental regulation of biosynthesis of a heterocyst envelope polysaccharide. Recombinant N-hexahistidine-tagged HepK (H6HepK), the cytoplasmic portion H6'HepK of H6HepK, H6DevR, and H6DevRA were overexpressed in Escherichia coli and purified to homogeneity. H6'HepK, but not H6HepK, autophosphorylates with [gamma-32P]ATP. ADP, specifically, elicits dephosphorylation of phosphorylated H6'HepK. The phosphoryl group of H6'HepK is transferred rapidly and efficiently to both H6DevR and H6DevRA but not to His-tagged OmpR, whose cognate sensor kinase is EnvZ. Sequence comparisons, the results of site-specific mutagenesis, and tests of chemical stability support identification of HepK-His348 and DevR-Asp53 as the phosphorylated residues. The mutation HepK-H348A abolishes both in vitro autokinase activity and in vivo functionality of HepK. Heterocysts of both hepK Anabaena and devRA Anabaena lack an envelope polysaccharide layer and are nonfunctional. Consistent with the normal site of deposition of that polysaccharide, a hepK::gfp transcriptional fusion is expressed principally in proheterocysts. HepK/DevRA is the first two-component system identified that regulates the biosynthesis of a polysaccharide as part of a patterned differentiation process.
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PMID:A two-component system mediates developmental regulation of biosynthesis of a heterocyst polysaccharide. 1263 41

The rhizobial FixL/FixJ system, a member of the superfamily of bacterial two-component signal transducing systems, regulates the expression of nitrogen fixation-related genes by sensing environmental oxygen tension. Oxygen-free (deoxy) FixL is autophosphorylated at an invariant histidine residue with ATP, and the phosphoryl group is transferred to FixJ, leading to an enhancement in transcriptional activity at low oxygen tensions, but the histidine kinase activity of the oxygen-bound (oxy) form is inhibited. To investigate the mechanism of oxygen sensing, we established a FixL/FixJ-mediated PfixK-lacZ reporter system in Escherichia coli, and isolated FixL and FixJ mutations conferring an upregulation of lacZ gene expression on the reporter cells even under aerobic conditions. FixL mutant proteins, which contain single amino acid changes near the autophosphorylation site, showed elevated levels of autophosphorylation and a concomitant phosphoryl transfer to FixJ in the presence of oxygen, although their oxygen-binding affinities were unimpaired. These mutational analyses suggest that the autophosphorylation domain plays a crucial role in regulatory coupling between oxygen binding and kinase activity. FixJ mutants in helix alpha1 and strand beta5 of the N-terminal half exhibited the formation of a stable acyl phosphate bond. In contrast, those in helices alpha4 and alpha5 constitutively bound to the fixK promoter in a monomeric form, suggesting that the alpha4 and alpha5 helices may be involved in the post-phosphorylation/dimerization signal transfer to liberate the DNA-binding activity of the C-terminal domain, not only serving as a dimerization interface.
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PMID:The uncoupling of oxygen sensing, phosphorylation signalling and transcriptional activation in oxygen sensor FixL and FixJ mutants. 1267 98

EnvZ, a dimeric transmembrane histidine kinase, belongs to the family of His-Asp phosphorelay signal transduction systems. The cytoplasmic kinase domain of EnvZ can be dissected into two independently functioning domains, A and B, whose NMR solution structures have been individually determined. Here, we examined the topological arrangement of these two domains in the EnvZ dimer, a structure that is key to understanding the mechanism underlying the autophosphorylation activity of the kinase. A series of cysteine substitution mutants were constructed to test the feasibility of chemical crosslinking between the two domains. These crosslinking data demonstrate that helix I of domain A of one subunit in the EnvZc dimer is in close proximity to domain B of the other subunit in the same dimer, while helix II of domain A of one subunit interacts with domain B of the same subunit in the EnvZc dimer. This is the first demonstration of the topological arrangement between the central dimerization domain containing the active center His residues (domain A) and the ATP-binding catalysis assisting domain (domain B) in a class I histidine kinase.
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PMID:Probing catalytically essential domain orientation in histidine kinase EnvZ by targeted disulfide crosslinking. 1269 49

The EnvZ/OmpR histidyl-aspartyl phosphorelay (HAP) system in Escherichia coli regulates the expression of ompF and ompC, the major outer membrane porin genes, in response to environmental osmolarity changes. Here, we report that dimers of EnvZc, the cytoplasmic domain of EnvZ, undergo spontaneous subunit exchange in solution. By introducing a cysteine substitution (S260C) in the dimerization domain of EnvZc, we were able to crosslink the two subunits in a dimer and trap the heterodimer formed between two different mutant EnvZc. By using a complementing system with two autophosphorylation-defective EnvZc mutants, one containing the H243V mutation at the autophosphorylation site and the other containing the G405A mutation in the ATP-binding domain, we demonstrated that an EnvZc(G405A) subunit can be phosphorylated by an EnvZc(H243V) subunit only when a heterodimer is formed. The rate of subunit exchange is concentration-dependent, with higher rates at higher concentrations of protein. The disulfide-crosslinked EnvZc(G405A) homodimer could not be phosphorylated by EnvZc(H243V), since the heterodimer formation between the two mutant proteins was blocked, indicating that autophosphorylation cannot occur by dimer-dimer interaction. By using MBP-deltaL-EnvZc(S260C) fusion protein (deltaL: the linker region, spanning residues 180-222, was deleted), it was found that in the disulfide-crosslinked MBP-deltaL-EnvZc(S260C)/deltaL-EnvZc(S260C/G405A) heterodimer, only the deltaL-EnvZc(S260C/G405A) subunit was phosphorylated but not the MBP-deltaL-EnvZc(S260C) subunit. Together, the present results provide biochemical evidence that EnvZ autophosphorylation occurs in trans and only within an EnvZ dimer.
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PMID:Spontaneous subunit exchange and biochemical evidence for trans-autophosphorylation in a dimer of Escherichia coli histidine kinase (EnvZ). 1276 31


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