1P229 Control of bacterial flagellar rotation via crosstalk from a non-cognate histidine kinase to the response regulator CheY(14. Chemoreception,Poster)

ABSTRACT In a two-component regulatory system, an important means of signal transduction in microorganisms, a sensor kinase phosphorylates a response regulator protein on an aspartyl residue, resulting in activation. The active site of the response regulator is highly charged (containing a lysine, the phosphorylatable aspartate, two additional aspartates involved in metal binding, and an Mg2+ ion), and introduction of the dianionic phosphoryl group results in the repositioning of charged moieties. Furthermore, substitution of one of the Mg2+-coordinating aspartates with lysine or arginine in the Escherichia coli chemotaxis response regulator CheY results in phosphorylation-independent activation. In order to examine the consequences of altered charge distribution for response regulator activity and to identify possible additional amino acid substitutions that result in phosphorylation-independent activation, we made 61 CheY mutants in which residues close to the site of phosphorylation (Asp57) were replaced by various charged amino acids. Most substitutions (47 of 61) resulted in the complete loss of CheY activity, as measured by the inability to support clockwise flagellar rotation. However, 10 substitutions, all introducing a new positive charge, resulted in the loss of chemotaxis but in the retention of some clockwise flagellar rotation. Of the mutants in this set, only the previously identified CheY13DK and CheY13DR mutants displayed clockwise activity in the absence of the CheA sensor kinase. The absence of negatively charged substitution mutants with residual activity suggests that the introduction of additional negative charges into the active site is particularly deleterious for CheY function. Finally, the spatial distribution of positions at which amino acid substitutions are functionally tolerated or not tolerated is consistent with the presently accepted mechanism of response regulator activation and further suggests a possible role for Met17 in signal transduction by CheY.

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The orphan histidine kinase PdtaS-p regulates both morphological differentiation and antibiotic biosynthesis together with the orphan response regulator PdtaR-p in Streptomyces

Microbiological Research ◽

10.1016/j.micres.2020.126411 ◽

2020 ◽

Vol 233 ◽

pp. 126411 ◽

Cited By ~ 2

Author(s):

Lei Li ◽

Yawei Zhao ◽

Jingjing Ma ◽

Hengnuo Tao ◽

Guosong Zheng ◽

...

Keyword(s):

Histidine Kinase ◽

Response Regulator ◽

Morphological Differentiation ◽

Antibiotic Biosynthesis

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Insights into histidine kinase activation mechanisms from the monomeric blue light sensor EL346

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1813586116 ◽

2019 ◽

Vol 116 (11) ◽

pp. 4963-4972 ◽

Cited By ~ 5

Author(s):

Igor Dikiy ◽

Uthama R. Edupuganti ◽

Rinat R. Abzalimov ◽

Peter P. Borbat ◽

Madhur Srivastava ◽

...

Keyword(s):

Blue Light ◽

Conformational Changes ◽

Histidine Kinase ◽

Structural Changes ◽

Molecular Mechanisms ◽

Catalytic Domain ◽

Response Regulator ◽

Phosphoryl Group ◽

Dark State ◽

Light Sensing

Translation of environmental cues into cellular behavior is a necessary process in all forms of life. In bacteria, this process frequently involves two-component systems in which a sensor histidine kinase (HK) autophosphorylates in response to a stimulus before subsequently transferring the phosphoryl group to a response regulator that controls downstream effectors. Many details of the molecular mechanisms of HK activation are still unclear due to complications associated with the multiple signaling states of these large, multidomain proteins. To address these challenges, we combined complementary solution biophysical approaches to examine the conformational changes upon activation of a minimal, blue-light–sensing histidine kinase from Erythrobacter litoralis HTCC2594, EL346. Our data show that multiple conformations coexist in the dark state of EL346 in solution, which may explain the enzyme’s residual dark-state activity. We also observe that activation involves destabilization of the helices in the dimerization and histidine phosphotransfer-like domain, where the phosphoacceptor histidine resides, and their interactions with the catalytic domain. Similar light-induced changes occur to some extent even in constitutively active or inactive mutants, showing that light sensing can be decoupled from activation of kinase activity. These structural changes mirror those inferred by comparing X-ray crystal structures of inactive and active HK fragments, suggesting that they are at the core of conformational changes leading to HK activation. More broadly, our findings uncover surprising complexity in this simple system and allow us to outline a mechanism of the multiple steps of HK activation.

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Kinetic Analysis of a Globin-Coupled Histidine Kinase, AfGcHK: Effects of the Heme Iron Complex, Response Regulator, and Metal Cations on Autophosphorylation Activity

Biochemistry ◽

10.1021/acs.biochem.5b00517 ◽

2015 ◽

Vol 54 (32) ◽

pp. 5017-5029 ◽

Cited By ~ 10

Author(s):

Veronika Fojtikova ◽

Martin Stranava ◽

Marten H. Vos ◽

Ursula Liebl ◽

Jakub Hranicek ◽

...

Keyword(s):

Kinetic Analysis ◽

Histidine Kinase ◽

Response Regulator ◽

Metal Cations ◽

Iron Complex ◽

Heme Iron ◽

Complex Response

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Identification of genes required for different stages of dendritic swarming in Bacillus subtilis, with a novel role for phrC

Microbiology ◽

10.1099/mic.0.021477-0 ◽

2009 ◽

Vol 155 (2) ◽

pp. 398-412 ◽

Cited By ~ 23

Author(s):

Kassem Hamze ◽

Daria Julkowska ◽

Sabine Autret ◽

Krzysztof Hinc ◽

Krzysztofa Nagorska ◽

...

Keyword(s):

Histidine Kinase ◽

Cytoplasmic Membrane ◽

Mutational Analysis ◽

Response Regulator ◽

Liquid Cultures ◽

Global Regulators ◽

Two Component ◽

Control Production ◽

Surfactin Production ◽

First Time

Highly branched dendritic swarming of B. subtilis on synthetic B-medium involves a developmental-like process that is absolutely dependent on flagella and surfactin secretion. In order to identify new swarming genes, we targeted the two-component ComPA signalling pathway and associated global regulators. In liquid cultures, the histidine kinase ComP, and the response regulator ComA, respond to secreted pheromones ComX and CSF (encoded by phrC) in order to control production of surfactin synthases and ComS (competence regulator). In this study, for what is believed to be the first time, we established that distinct early stages of dendritic swarming can be clearly defined, and that they are amenable to genetic analysis. In a mutational analysis producing several mutants with distinctive phenotypes, we were able to assign the genes sfp (activation of surfactin synthases), comA, abrB and codY (global regulators), hag (flagellin), mecA and yvzB (hag-like), and swrB (motility), to the different swarming stages. Surprisingly, mutations in genes comPX, comQ, comS, rapC and oppD, which are normally indispensable for import of CSF, had only modest effects, if any, on swarming and surfactin production. Therefore, during dendritic swarming, surfactin synthesis is apparently subject to novel regulation that is largely independent of the ComXP pathway; we discuss possible alternative mechanisms for driving srfABCD transcription. We showed that the phrC mutant, largely independent of any effect on surfactin production, was also, nevertheless, blocked early in swarming, forming stunted dendrites, with abnormal dendrite initiation morphology. In a mixed swarm co-inoculated with phrC sfp+ and phrC+ sfp (GFP), an apparently normal swarm was produced. In fact, while initiation of all dendrites was of the abnormal phrC type, these were predominantly populated by sfp cells, which migrated faster than the phrC cells. This and other results indicated a specific migration defect in the phrC mutant that could not be trans-complemented by CSF in a mixed swarm. CSF is the C-terminal pentapeptide of the surface-exposed PhrC pre-peptide and we propose that the residual PhrC 35 aa residue peptide anchored in the exterior of the cytoplasmic membrane has an apparently novel extracellular role in swarming.

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Functional reconstitution of the Salmonella typhimurium PhoQ histidine kinase sensor in proteoliposomes

Biochemical Journal ◽

10.1042/bj20050060 ◽

2005 ◽

Vol 390 (3) ◽

pp. 769-776 ◽

Cited By ~ 19

Author(s):

Sarah Sanowar ◽

Hervé Le Moual

Keyword(s):

Salmonella Typhimurium ◽

Histidine Kinase ◽

Catalytic Domain ◽

Response Regulator ◽

Phosphoryl Group ◽

Two Component ◽

Two Component Signal Transduction ◽

High Concentrations

Two-component signal-transduction systems are widespread in bacteria. They are usually composed of a transmembrane histidine kinase sensor and a cytoplasmic response regulator. The PhoP/PhoQ two-component system of Salmonella typhimurium contributes to virulence by co-ordinating the adaptation to low concentrations of environmental Mg2+. Limiting concentrations of extracellular Mg2+ activate the PhoP/PhoQ phosphorylation cascade modulating the transcription of PhoP-regulated genes. In contrast, high concentrations of extracellular Mg2+ stimulate the dephosphorylation of the response regulator PhoP by the PhoQ kinase sensor. In the present study, we report the purification and functional reconstitution of PhoQHis, a PhoQ variant with a C-terminal His tag, into Escherichia coli liposomes. The functionality of PhoQHis was essentially similar to that of PhoQ as shown in vivo and in vitro. Purified PhoQHis was inserted into liposomes in a unidirectional orientation, with the sensory domain facing the lumen and the catalytic domain facing the extraluminal environment. Reconstituted PhoQHis exhibited all the catalytic activities that have been described for histidine kinase sensors. Reconstituted PhoQHis was capable of autokinase activity when incubated in the presence of Mg2+-ATP. The phosphoryl group could be transferred from reconstituted PhoQHis to PhoP. Reconstituted PhoQHis catalysed the dephosphorylation of phospho-PhoP and this activity was stimulated by the addition of extraluminal ADP.

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A Novel Redox-Sensing Histidine Kinase That Controls Carbon Catabolite Repression inAzoarcussp. CIB

mBio ◽

10.1128/mbio.00059-19 ◽

2019 ◽

Vol 10 (2) ◽

Author(s):

J. Andrés Valderrama ◽

Helena Gómez-Álvarez ◽

Zaira Martín-Moldes ◽

M. Álvaro Berbís ◽

F. Javier Cañada ◽

...

Keyword(s):

Aromatic Compounds ◽

Histidine Kinase ◽

Catabolite Repression ◽

Carbon Catabolite Repression ◽

Response Regulator ◽

Sensor Kinase ◽

General Role ◽

Two Component ◽

Redox Switch ◽

Redox Sensing

ABSTRACTWe have identified and characterized the AccS multidomain sensor kinase that mediates the activation of the AccR master regulator involved in carbon catabolite repression (CCR) of the anaerobic catabolism of aromatic compounds inAzoarcussp. CIB. A truncated AccS protein that contains only the soluble C-terminal autokinase module (AccS′) accounts for the succinate-dependent CCR control. In vitroassays with purified AccS′ revealed its autophosphorylation, phosphotransfer from AccS′∼P to the Asp60 residue of AccR, and the phosphatase activity toward its phosphorylated response regulator, indicating that the equilibrium between the kinase and phosphatase activities of AccS′ may control the phosphorylation state of the AccR transcriptional regulator. Oxidized quinones, e.g., ubiquinone 0 and menadione, switched the AccS′ autokinase activity off, and three conserved Cys residues, which are not essential for catalysis, are involved in such inhibition. Thiol oxidation by quinones caused a change in the oligomeric state of the AccS′ dimer resulting in the formation of an inactive monomer. This thiol-based redox switch is tuned by the cellular energy state, which can change depending on the carbon source that the cells are using. This work expands the functional diversity of redox-sensitive sensor kinases, showing that they can control new bacterial processes such as CCR of the anaerobic catabolism of aromatic compounds. The AccSR two-component system is conserved in the genomes of some betaproteobacteria, where it might play a more general role in controlling the global metabolic state according to carbon availability.IMPORTANCETwo-component signal transduction systems comprise a sensor histidine kinase and its cognate response regulator, and some have evolved to sense and convert redox signals into regulatory outputs that allow bacteria to adapt to the altered redox environment. The work presented here expands knowledge of the functional diversity of redox-sensing kinases to control carbon catabolite repression (CCR), a phenomenon that allows the selective assimilation of a preferred compound among a mixture of several carbon sources. The newly characterized AccS sensor kinase is responsible for the phosphorylation and activation of the AccR master regulator involved in CCR of the anaerobic degradation of aromatic compounds in the betaproteobacteriumAzoarcussp. CIB. AccS seems to have a thiol-based redox switch that is modulated by the redox state of the quinone pool. The AccSR system is conserved in several betaproteobacteria, where it might play a more general role controlling their global metabolic state.

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The Hybrid Histidine Kinase Hk1 Is Part of a Two-Component System That Is Essential for Survival of Borrelia burgdorferi in Feeding Ixodes scapularis Ticks

Infection and Immunity ◽

10.1128/iai.05136-11 ◽

2011 ◽

Vol 79 (8) ◽

pp. 3117-3130 ◽

Cited By ~ 52

Author(s):

Melissa J. Caimano ◽

Melisha R. Kenedy ◽

Toru Kairu ◽

Daniel C. Desrosiers ◽

Michael Harman ◽

...

Keyword(s):

Borrelia Burgdorferi ◽

Histidine Kinase ◽

Ixodes Scapularis ◽

Response Regulator ◽

Content Type ◽

Diguanylate Cyclases ◽

Two Component ◽

Tick Transmission ◽

Two Component Systems ◽

Hybrid Histidine Kinase

ABSTRACTTwo-component systems (TCS) are principal mechanisms by which bacteria adapt to their surroundings.Borrelia burgdorferiencodes only two TCS. One is comprised of a histidine kinase, Hk2, and the response regulator Rrp2. While the contribution of Hk2 remains unclear, Rrp2 is part of a regulatory pathway involving the spirochete's alternate sigma factors, RpoN and RpoS. Genes within the Rrp2/RpoN/RpoS regulon function to promote tick transmission and early infection. The other TCS consists of a hybrid histidine kinase, Hk1, and the response regulator Rrp1. Hk1 is composed of two periplasmic sensor domains (D1 and D2), followed by conserved cytoplasmic histidine kinase core, REC, and Hpt domains. In addition to its REC domain, Rrp1 contains a GGDEF motif characteristic of diguanylate cyclases. To investigate the role of Hk1 during the enzootic cycle, we inactivated this gene in two virulent backgrounds. Extensive characterization of the resulting mutants revealed a dramatic phenotype whereby Hk1-deficient spirochetes are virulent in mice and able to migrate out of the bite site during feeding but are killed within the midgut following acquisition. We hypothesize that the phosphorelay between Hk1 and Rrp1 is initiated by the binding of feeding-specific ligand(s) to Hk1 sensor domain D1 and/or D2. Once activated, Rrp1 directs the synthesis of cyclic dimeric GMP (c-di-GMP), which, in turn, modulates the expression and/or activity of gene products required for survival within feeding ticks. In contrast to the Rrp2/RpoN/RpoS pathway, which is active only within feeding nymphs, the Hk1/Rrp1 TCS is essential for survival during both larval and nymphal blood meals.

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