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)

CheA is a histidine kinase central to the signal transduction pathway for chemotaxis in Escherichia coli. CheA autophosphorylates at His-48, with ATP as the phosphodonor, and then donates its phosphoryl groups to two aspartate autokinases, CheY and CheB. Phospho-CheY controls the flagellar motors, whereas phospho-CheB participates in sensory adaptation. Polypeptides encompassing the N-terminal P1 domain of CheA can be transphosphorylated in vitro by the CheA catalytic domain and yet have no deleterious effect on chemotactic ability when expressed at high levels in wild-type cells. To find out why, we examined the effects of a purified P1 fragment, CheA[1-149], on CheA-related signaling activities in vitro and devised in vivo assays for those same activities. Although readily phosphorylated by CheA[260-537], the CheA catalytic domain, CheA[1-149], was a poor substrate for transphosphorylation by full-length CheA molecules, implying that the resident P1 domain monopolizes the CheA catalytic center. CheA-H48Q, a nonphosphorylatable mutant, failed to transphosphorylate CheA[1-149], suggesting that phosphorylation of the P1 domain in cis may alleviate the exclusion effect. In agreement with these findings, a 40-fold excess of CheA[1-149] fragments did not impair the CheA autophosphorylation reaction. CheA[1-149] did acquire phosphoryl groups via reversible phosphotransfer reactions with CheB and CheY molecules. An H48Q mutant of CheA[1-149] could not participate in these reactions, indicating that His-48 is probably the substrate site. The low level of efficiency of these phosphotransfer reactions and the inability of CheA[1-149] to interfere with CheA autophosphorylation most likely account for the failure of liberated P1 domains to jam chemotactic signaling in wild-type cells. However, an excess of CheA[1-149] fragments was able to support chemotactic signaling by P1-deficient cheA mutants, demonstrating that CheA[1-149] fragments have both transphosphorylation and phosphotransfer capability in vivo.
J Bacteriol 1996 Dec
PMID:Chemotactic signaling by the P1 phosphorylation domain liberated from the CheA histidine kinase of Escherichia coli. 895 92

Two-component regulatory systems require highly specific interactions between histidine kinase (transmitter) and response regulator (receiver) proteins. We have developed a novel genetic strategy that is based on tightly regulated synthesis of a given protein to identify domains and residues of an interacting protein that are critical for interactions between them. Using a reporter strain synthesizing the nonpartner kinase VanS under tight arabinose control and carrying a promoter-lacZ fusion activated by phospho-PhoB, we isolated altered recognition (AR) mutants of PhoB showing enhanced activation (phosphorylation) by VanS as arabinose-dependent Lac+ mutants. Changes in the PhoBAR mutants cluster in a "patch" near the proposed helix 4 of PhoB based on the CheY crystal structure (a homolog of the PhoB receiver domain) providing further evidence that helix 4 lies in the kinase-regulator interface. Based on the CheY structure, one mutant has an additional change in a region that may propagate a conformational change to helix 4. The overall genetic strategy described here may also be useful for studying interactions of other components of the vancomycin resistance and P1 signal transduction pathways, other two-component regulatory systems, and other interacting proteins. Conditionally replicative oriRR6K gamma attP "genome targeting" suicide plasmids carrying mutagenized phoB coding regions were integrated into the chromosome of a reporter strain to create mutant libraries; plasmids encoding mutant PhoB proteins were subsequently retrieved by P1-Int-Xis cloning. Finally, the use of similar genome targeting plasmids and P1-Int-Xis cloning should be generally useful for constructing genomic libraries from a wide array of organisms.
Proc Natl Acad Sci U S A 1996 Dec 10
PMID:Altered recognition mutants of the response regulator PhoB: a new genetic strategy for studying protein-protein interactions. 896 56

Keratins are the major structural proteins of keratinocytes, which are the most abundant cell type in the mammalian epidermis. Mutations in epidermal keratin genes have been shown to cause severe blistering skin abnormalities. One such disease, epidermolytic hyperkeratosis (EHK), also known as bullous congenital ichthyosiform erythroderma, occurs as a result of mutations in highly conserved regions of keratins K1 and K10. Patients with EHK first exhibit erythroderma with severe blistering, which later is replaced by thick patches of scaly skin. To assess the effect of a mutated K1 gene on skin biology and to produce an animal model for EHK, we removed 60 residues from the 2B segment of HK1 and observed the effects of its expression in the epidermis of transgenic mice. Phenotypes of the resultant mice closely resembled those observed in the human disease, first with epidermal blisters, then later with hyperkeratotic lesions. In neonatal mice homozygous for the transgene, the skin was thicker, with an increased labeling index, and the spinous cells showed a collapse of the keratin filament network around the nuclei, suggesting that a critical concentration of the mutant HK1, over the endogenous MK1, was required to disrupt the structural integrity of the spinous cells. Additionally, footpad epithelium, which is devoid of hair follicles, showed blistering in the spinous layer, suggesting that hair follicles can stabilize or protect the epidermis from trauma. Blisters were not evident in adult mice, but instead they showed a thick, scaly hyperkeratotic skin with increased mitosis, resulting in an increased number of corneocytes and granular cells. Irregularly shaped keratohyalin granules were also observed. To date, this is the only transgenic model to show the typical morphology found in the adult form of EHK.
Differentiation 1996 Dec
PMID:A transgenic mouse model that recapitulates the clinical features of both neonatal and adult forms of the skin disease epidermolytic hyperkeratosis. 898 79

Cells react to increased osmolality with numerous changes in gene expression. The specific genes affected differ between species, but the known osmoprotective effects of the gene products are remarkably similar, particularly with regard to cellular accumulation of compatible organic osmolytes. Here we concentrate on the molecular basis for osmotic regulation of gene expression, emphasizing certain genes expressed in bacteria, yeast, and the mammalian renal medulla because their expression is best understood. Thus, we emphasize 1) bacterial and yeast two-component histidine kinase systems, each consisting of a membrane osmolality sensor and a separate cytoplasmic response regulator that, when phosphorylated, alters transcription, 2) volume regulatory increases in cellular K+ salts that can prompt increased gene transcription in bacteria through direct effects on DNA and that in mammalian renal cells increase transcription, seemingly via trans-activating proteins, 3) a yeast kinase cascade that transmits an osmotic signal to the gene regulating the level of glycerol, and 4) in mammalian cells, several homologous cascades that are activated by hypertonicity, but whose osmoregulatory targets are not yet known.
FASEB J 1996 Dec
PMID:Osmotic regulation of gene expression. 900 51

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.
J Bacteriol 1997 Dec
PMID:Transduction of envelope stress in Escherichia coli by the Cpx two-component system. 940 Oct 31

The transmembrane, homodimeric aspartate receptor of Escherichia coli and Salmonella typhimurium controls the chemotactic response to aspartate, an attractant, by regulating the activity of a cytoplasmic histidine kinase. The cytoplasmic domain of the receptor plays a central role in both kinase regulation and sensory adaptation, although its structure and regulatory mechanisms are unknown. The present study utilizes cysteine and disulfide scanning to probe residues Leu-250 through Gln-309, a region that contains the first of two adaptive methylation segments within the cytoplasmic domain. Following the introduction of consecutive cysteine residues by scanning mutagenesis, the measurement of sulfhydryl chemical reactivities reveals an alpha-helical pattern of exposed and buried positions spanning residues 270-309. This detected helix, termed the "first methylation helix," is strongly amphiphilic; its exposed face is highly anionic and possesses three methylation sites, while its buried face is hydrophobic. In vivo and in vitro assays of receptor function indicate that inhibitory cysteine substitutions are most prevalent on the buried face of the first methylation helix, demonstrating that this face is involved in a critical packing interaction. The buried face is further analyzed by disulfide scanning, which reveals three "lock-on" disulfides that covalently trap the receptor in its kinase-activating state. Each of the lock-on disulfides cross-links the buried faces of the two symmetric first methylation helices of the dimer, placing these helices in direct contact at the subunit interface. Comparative sequence analysis of 56 related receptors suggests that the identified helix is a conserved feature of this large receptor family, wherein it is likely to play a general role in adaptation and kinase regulation. Interestingly, the rapid rates and promiscuous nature of disulfide formation reactions within the scanned region reveal that the cytoplasmic domain of the full-length, membrane-bound receptor has a highly dynamic structure. Overall, the results demonstrate that cysteine and disulfide scanning can identify secondary structure elements and functionally important packing interfaces, even in proteins that are inaccessible to other structural methods.
J Biol Chem 1997 Dec 26
PMID:Cysteine and disulfide scanning reveals a regulatory alpha-helix in the cytoplasmic domain of the aspartate receptor. 940 66

Bacterial chemotaxis is the most studied model system for signaling by the widely spread family of two-component regulatory systems. It is controlled by changes in the phosphorylation level of the chemotactic response regulator, CheY, mediated by a histidine kinase (CheA) and a specific phosphatase (CheZ). While it is known that CheA activity is regulated, via the receptors, by chemotactic stimuli, the input that may regulate CheY dephosphorylation by CheZ has not been found. We measured, by using stopped-flow fluorometry, the kinetics of CheZ-mediated dephosphorylation of CheY. The onset of dephosphorylation was delayed by approximately 50 ms after mixing phosphorylated CheY (CheY approximately P) with CheZ, and a distinct overshoot was observed in the approach to the new steady state of CheY approximately P. The delay and overshoot were not observed in a hyperactive mutant CheZ protein (CheZ54RC) that does not support chemotaxis in vivo and appears to be constitutively active. CheZ activity was cooperative with respect to CheY approximately P, with a Hill-coefficient of 2.5. The observed delayed modulation of CheZ activity and its cooperativity suggest that the phosphatase activity is regulated at the level of CheY approximately P-CheZ interaction. This novel kind of interplay between a response regulator and its phosphatase may be involved in signal tuning and in adaptation to chemotactic signals.
J Mol Biol 1998 Dec 11
PMID:Regulation of phosphatase activity in bacterial chemotaxis. 983 37

The Saccharomyces cerevisiae Sln1 protein is a 'two-component' regulator involved in osmotolerance. Two-component regulators are a family of signal-transduction molecules with histidine kinase activity common in prokaryotes and recently identified in eukaryotes. Phosphorylation of Sln1p inhibits the HOG1 MAP kinase osmosensing pathway via a phosphorelay mechanism including Ypd1p and the response regulator, Ssk1p. SLN1 also activates an MCM1-dependent reporter gene, P-lacZ, but this function is independent of Ssk1p. We present genetic and biochemical evidence that Skn7p is the response regulator for this alternative Sln1p signaling pathway. Thus, the yeast Sln1 phosphorelay is actually more complex than appreciated previously; the Sln1 kinase and Ypd1 phosphorelay intermediate regulate the activity of two distinct response regulators, Ssk1p and Skn7p. The established role of Skn7p in oxidative stress is independent of the conserved receiver domain aspartate, D427. In contrast, we show that Sln1p activation of Skn7p requires phosphorylation of D427. The expression of TRX2, previously shown to exhibit Skn7p-dependent oxidative-stress activation, is also regulated by the SLN1 phosphorelay functions of Skn7p. The identification of genes responsive to both classes of Skn7p function suggests a central role for Skn7p and the SLN1-SKN7 pathway in integrating and coordinating cellular response to various types of environmental stress.
EMBO J 1998 Dec 01
PMID:The yeast histidine protein kinase, Sln1p, mediates phosphotransfer to two response regulators, Ssk1p and Skn7p. 984 1

Recent evidence suggests that cholesterol sulfate may be an important second messenger involved in signaling epidermal differentiation in skin. The activity of cholesterol sulfotransferase (Ch-ST) is increased during squamous differentiation of keratinocytes and is believed to be a marker enzyme for terminal differentiation. The primary objective of this study was to examine changes in levels of cholesterol sulfate (CS) and activity of its biosynthetic enzyme, Ch-ST, during multistage carcinogenesis in mouse skin. Using SENCAR mice, we determined the activity of Ch-ST in normal epidermis, in tumor promoter-treated epidermis, in epidermis during wound healing, and in mouse skin tumors generated by initiation-promotion regimens. A single topical application of tumor promoters led to significantly elevated levels of Ch-ST activity and of CS. Epidermal Ch-ST activity was also elevated during wound healing. Dramatic increases in CS levels and in the activity of Ch-ST were found in nearly all of the papillomas and squamous cell carcinomas examined. The increased levels of CS and activity of Ch-ST in tumor promoter-treated epidermis were accompanied by increased transglutaminase-I activity. In contrast, transglutaminase I activity was not elevated in primary papillomas or squamous cell carcinomas. Finally, Ch-ST activity was significantly elevated in the epidermis of newborn HK1.ras transgenic mice, whereas transglutaminase I activity did not correlate with Ch-ST activity in these mice. These results demonstrate that diverse tumor-promoting stimuli all produce elevated CS levels and Ch-ST activity and that CS levels and Ch-ST activity were constitutively elevated in both papillomas and squamous cell carcinomas. The data also suggest a mechanism for upregulation of Ch-ST in skin tumors involving activation/upregulation of Ha-ras.
J Invest Dermatol 1998 Dec
PMID:Alterations in cholesterol sulfate and its biosynthetic enzyme during multistage carcinogenesis in mouse skin. 985 4

The FixL proteins are biological oxygen sensors that restrict the expression of specific genes to hypoxic conditions. FixL's oxygen-detecting domain is a heme binding region that controls the activity of an attached histidine kinase. The FixL switch is regulated by binding of oxygen and other strong-field ligands. In the absence of bound ligand, the heme domain permits kinase activity. In the presence of bound ligand, this domain turns off kinase activity. Comparison of the structures of two forms of the Bradyrhizobium japonicum FixL heme domain, one in the "on" state without bound ligand and one in the "off" state with bound cyanide, reveals a mechanism of regulation by a heme that is distinct from the classical hemoglobin models. The close structural resemblance of the FixL heme domain to the photoactive yellow protein confirms the existence of a PAS structural motif but reveals the presence of an alternative regulatory gateway.
Proc Natl Acad Sci U S A 1998 Dec 22
PMID:Structure of a biological oxygen sensor: a new mechanism for heme-driven signal transduction. 986 Sep 42


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