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)

Previous studies described lacZ'- and cat'-'cpxA fusion genes whose expression restored to normal all the phenotypic defects associated with cpxA mutations (Albin, R., and Silverman, P. M. (1984) Mol. Gen. Genet. 197, 272-279). Here, we show by DNA nucleotide sequence analysis that the fusion genes encode 241 carboxyl-terminal amino acids of the CpxA polypeptide. Using this information, we constructed a fusion gene containing the same 241 cpxA codons preceded by 1007 codons of beta-galactosidase. The resultant hybrid polypeptide was purified and used to raise an anti-(CpxA polypeptide) antiserum. Using the antiserum, we have identified the chromsomal Escherichia coli K12 cpxA gene product as a 52-kDa polypeptide. The polypeptide showed temperature-sensitive accumulation in a strain carrying both the cpxA2[Ts] and cpxB1 alleles and accumulated to a level higher than normal in cells that carried a high-copy number, cpxA+ plasmid. Immune precipitates of in vitro transcription-translation reactions with cpxA+ plasmids as template also contained a 52-kDa polypeptide, indistinguishable in electrophoretic mobility from the immunoreactive polypeptide synthesized in vivo. Two regions of amino acid sequence at the carboxyl-terminus of the CpxA polypeptide are significantly homologous to corresponding regions of the E. coli K12 EnvZ polypeptide, an inner membrane component that, like the CpxA polypeptide, is required to maintain the protein composition of the cell envelope. The cpxA coding sequence is followed by two repetitive extragenic palindrome sequences in opposite orientation.
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PMID:The Cpx proteins of Escherichia coli K12. Immunologic detection of the chromosomal cpxA gene product. 300 73

Specific induction of the copper resistance operon (cop) promoter from Pseudomonas syringae was measured by beta-galactosidase production from a cop promoter-lacZ fusion. Induction of the cop promoter in P. syringae pv. syringae required trans-acting factors from copper resistance plasmid pPT23D, from which cop was originally cloned. Tn5 mutagenesis of pPT23D was used to localize two complementation groups immediately downstream from copABCD. Cloning and sequencing of the DNA in this region revealed two genes, copR and copS, expressed in the same orientation as the cop operon but from a separate constitutive promoter. The amino acid sequence deduced from these genes showed distinct similarities to known two-component regulatory systems, including PhoB-PhoR and OmpR-EnvZ. In addition, CopR showed strong similarity to copper resistance activator protein PcoR from Escherichia coli. Functional chromosomal homologs to copRS activated the cop promoter, in a copper-inducible manner, in copper-resistant or -sensitive strains of P. syringae pv. tomato and other Pseudomonas species. This implies that copper-inducible gene regulation is associated with a common chromosomally encoded function, as well as plasmid-borne copper resistance, in Pseudomonas spp.
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PMID:A two-component regulatory system required for copper-inducible expression of the copper resistance operon of Pseudomonas syringae. 844 73

The histidine kinase/phosphatase EnvZ helps Escherichia coli adapt to osmotic shock by controlling the phosphorylation state of the transcription factor OmpR, which regulates the levels of the outer membrane porin proteins OmpF and OmpC. We examined the effects of mutating the highly conserved Thr(247) residue in EnvZ. Using purified C-terminal domains of wild-type and mutant EnvZ proteins, we demonstrate that Thr(247) plays a vital role in EnvZ function, variously affecting its autokinase and phosphotransferase activities, but mostly its function as a phosphatase. The cytoplasmic domain of EnvZ (EnvZc) is composed of three segments: the linker domain (residues 180-222), domain A (residues 223-289), and domain B (residues 290-450). It has been shown that the isolated domain A itself can dephosphorylate phosphorylated OmpR. Here we show that mutating Thr(247) to Arg in domain A abolishes its phosphatase activity. Furthermore, using an in vivo beta-galactosidase activity assay of Taz1-1 (hybrid of the aspartate receptor Tar and EnvZ) constructs of the Thr(247) mutants in RU1012 cells expressing ompC-lacZ, we demonstrate that the external signal primarily down-regulates the phosphatase activity of EnvZ. Of the nine EnvZc(T247X) mutants (X = Ser, Ala, Cys, Lys, Asn, Glu, Gln, Tyr, or Arg) analyzed, only Ser functionally substituted for Thr at this position, whereas all the others displayed constitutive expression of beta-galactosidase.
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PMID:The critical role of the conserved Thr247 residue in the functioning of the osmosensor EnvZ, a histidine Kinase/Phosphatase, in Escherichia coli. 1097 66

The TrcRS two-component system of Mycobacterium tuberculosis is comprised of the TrcS histidine kinase and the TrcR response regulator, which is homologous to the OmpR class of DNA binding response regulators. Reverse transcription-PCRs with total RNA showed that the trcR and trcS two-component system genes are transcribed in broth-grown M. tuberculosis. Analysis of the trcR and trcS genes using various SCOTS (selective capture of transcribed sequences) probes also confirmed that these genes are expressed in broth-grown cultures and after 18 h of M. tuberculosis growth in cultured human primary macrophages. To determine if the TrcR response regulator is autoregulated, a trcR-lacZ fusion plasmid and a TrcR expression plasmid were cotransformed into Escherichia coli. Upon induction of the TrcR protein, there was a >500-fold increase in beta-galactosidase activity from the trcR-lacZ fusion, indicating that TrcR is involved in transcriptional autoactivation. Gel mobility shift assays with the trcR promoter and TrcR established that the response regulator was autoregulating via direct binding. By use of a delimiting series of overlapping trcR PCR fragments in gel mobility shift assays with TrcR, an AT-rich region of the trcR promoter was shown to be essential for TrcR binding. Additionally, this AT-rich sequence was protected by TrcR in DNase I protection assays. To further analyze the role of the AT-rich region in TrcR autoregulation, the trcR promoter was mutated and analyzed in lacZ transcriptional fusions in the presence of TrcR. Alteration of the AT-rich sequence in the trcR promoter resulted in the loss of trcR transcriptional activation in the presence of TrcR. This report indicates that the M. tuberculosis TrcR response regulator activates its own expression by interacting with the AT-rich sequence of the trcR promoter.
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PMID:Expression, autoregulation, and DNA binding properties of the Mycobacterium tuberculosis TrcR response regulator. 1191 51

Natural competence for genetic transformation in Streptococcus pneumoniae is controlled by the ComCDE signal-transduction pathway. Together, ComD, a membrane histidine kinase, and ComE, its cognate response regulator, constitute a typical two-component regulatory system involved in sensing the comC-encoded competence-stimulating peptide (CSP). The comCDE operon is strongly upregulated when CSP reaches a critical threshold, probably to coordinate competence induction throughout the population. During a study of the early regulation of the comCDE operon, a mutation which resulted in increased beta-galactosidase production from a comC : : lacZ fusion was isolated. This mutation, which was characterized as a G-->T change in the transcription terminator of the tRNA(Arg) located immediately upstream of comCDE, is suggested to destabilize the terminator and to allow transcriptional readthrough of comCDE. Here, it is shown that, quite unexpectedly, the mutation confers reduced transformability. A series of experiments undertaken with the aim of understanding this surprising phenotype is described. Evidence is presented that increased basal-level expression of comDE impedes both spontaneous and CSP-induced competence in S. pneumoniae. There is a discussion of how an increased concentration of ComD and/or ComE could affect competence development.
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PMID:Inhibition of competence development in Streptococcus pneumoniae by increased basal-level expression of the ComDE two-component regulatory system. 1643 20

Production of the lantibiotic subtilin in Bacillus subtilis ATCC 6633 is regulated in a quorum sensing-like mechanism with subtilin acting as autoinducer and signal transduction via the subtilin-specific two-component regulation system SpaRK. Here, we report the construction and application of a subtilin reporter strain in which subtilin induced lacZ gene expression in a B. subtilis ATCC 6633 spaS gene deletion mutant is monitored and visualized by the beta-galactosidase in a chromogenic plate assay. A quantitative microtiter plate subtilin bioassay was developed and optimized. Maximal sensitivity of the system was achieved after 6 h of incubation of the reporter strain together with subtilin in a medium containing 300 mM NaCl. This sensitive and unsusceptible method was applied to identify subtilin producing B. subtilis wild type strains from both, culture collections and soil samples. The B. subtilis lantibiotic ericin S with four amino acid exchanges compared to subtilin induces the subtilin reporter strain, in contrast to the structurally closely related Lactococcus lactis lantibiotic nisin. These observations suggest a certain substrate specificity of the histidine kinase SpaK, which however, also would allow the identification of subtilin-isoform producing microorganisms.
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PMID:Development and application of a microtiter plate-based autoinduction bioassay for detection of the lantibiotic subtilin. 1753 72

The two-component system SaeRS of Staphylococcus aureus is closely involved in the regulation of major virulence factors. However, little is known about the signals leading to saeRS activation. A total of four overlapping transcripts (T1 to T4) from three different transcription starting points are expressed in the sae operon. We used a beta-galactosidase reporter assay to characterize the putative promoter regions within the saeRS upstream region. The main transcript T2 is probably generated by endoribonucleolytic processing of the T1 transcript. Only two distinct promoter elements (P1 and P3) could be detected within the saeRS upstream region. The P3 promoter, upstream of saeRS, generates the T3 transcript, includes a cis-acting enhancer element and is repressed by saeRS. The most distal P1 promoter is strongly autoregulated, activated by agr, and repressed by sigma factor B. In strain Newman a mutation within the histidine kinase SaeS leads to a constitutively activated sae system. Evaluation of different external signals revealed that the P1 promoter in strain ISP479R and strain UAMS-1 is inhibited by low pH and high NaCl concentrations but activated by hydrogen peroxide. The most prominent induction of P1 was observed at subinhibitory concentrations of alpha-defensins in various S. aureus strains, with the exception of strain ISP479R and strain COL. P1 was not activated by the antimicrobial peptides LL37 and daptomycin. In summary, the results indicate that the sensor molecule SaeS is activated by alteration within the membrane allowing the pathogen to react to phagocytosis related effector molecules.
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PMID:The virulence regulator Sae of Staphylococcus aureus: promoter activities and response to phagocytosis-related signals. 1834 60

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