scholarly journals Structures of full-length VanR from Streptomyces coelicolor in both the inactive and activated states

2021 ◽  
Vol 77 (8) ◽  
Author(s):  
Lina J. Maciunas ◽  
Nadia Porter ◽  
Paula J. Lee ◽  
Kushol Gupta ◽  
Patrick J. Loll
Keyword(s):  
Resistance Genes ◽  
Bacterial Infections ◽  
Analytical Ultracentrifugation ◽  
Response Regulator ◽  
Streptomyces Coelicolor ◽  
Full Length ◽  
Phosphoryl Group ◽  
Two Component System ◽  
Receiver Domain ◽  
Serious Bacterial Infections

Vancomycin has historically been used as a last-resort treatment for serious bacterial infections. However, vancomycin resistance has become widespread in certain pathogens, presenting a serious threat to public health. Resistance to vancomycin is conferred by a suite of resistance genes, the expression of which is controlled by the VanR–VanS two-component system. VanR is the response regulator in this system; in the presence of vancomycin, VanR accepts a phosphoryl group from VanS, thereby activating VanR as a transcription factor and inducing expression of the resistance genes. This paper presents the X-ray crystal structures of full-length VanR from Streptomyces coelicolor in both the inactive and activated states at resolutions of 2.3 and 2.0 Å, respectively. Comparison of the two structures illustrates that phosphorylation of VanR is accompanied by a disorder-to-order transition of helix 4, which lies within the receiver domain of the protein. This transition generates an interface that promotes dimerization of the receiver domain; dimerization in solution was verified using analytical ultracentrifugation. The inactive conformation of the protein does not appear intrinsically unable to bind DNA; rather, it is proposed that in the activated form DNA binding is enhanced by an avidity effect contributed by the receiver-domain dimerization.

scholarly journals Crystal Structure of the Response Regulator 02 Receiver Domain, the Essential YycF Two-Component System of Streptococcus pneumoniae in both Complexed and Native States

2004 ◽  
Vol 186 (9) ◽  
pp. 2872-2879 ◽  
Author(s):  
Colin J. Bent ◽  
Neil W. Isaacs ◽  
Timothy J. Mitchell ◽  
Alan Riboldi-Tunnicliffe
Keyword(s):  
Streptococcus Pneumoniae ◽  
Active Site ◽  
Response Regulator ◽  
Thermotoga Maritima ◽  
Two Component System ◽  
Environmental Signals ◽  
Receiver Domain ◽  
Two Component ◽  
Protein Sequence Homology ◽  
Two Component Signal Transduction

ABSTRACT A variety of bacterial cellular responses to environmental signals are mediated by two-component signal transduction systems comprising a membrane-associated histidine protein kinase and a cytoplasmic response regulator (RR), which interpret specific stimuli and produce a measured physiological response. In RR activation, transient phosphorylation of a highly conserved aspartic acid residue drives the conformation changes needed for full activation of the protein. Sequence homology reveals that RR02 from Streptococcus pneumoniae belongs to the OmpR subfamily of RRs. The structures of the receiver domains from four members of this family, DrrB and DrrD from Thermotoga maritima, PhoB from Escherichia coli, and PhoP from Bacillus subtilis, have been elucidated. These domains are globally very similar in that they are composed of a doubly wound α5β5; however, they differ remarkably in the fine detail of the β4-α4 and α4 regions. The structures presented here reveal a further difference of the geometry in this region. RR02 is has been shown to be the essential RR in the gram-positive bacterium S. pneumoniae R. Lange, C. Wagner, A. de Saizieu, N. Flint, J. Molnos, M. Stieger, P. Caspers, M. Kamber, W. Keck, and K. E. Amrein, Gene 237:223-234, 1999; J. P. Throup, K. K. Koretke, A. P. Bryant, K. A. Ingraham, A. F. Chalker, Y. Ge, A. Marra, N. G. Wallis, J. R. Brown, D. J. Holmes, M. Rosenberg, and M. K. Burnham, Mol. Microbiol. 35:566-576, 2000). RR02 functions as part of a phosphotransfer system that ultimately controls the levels of competence within the bacteria. Here we report the native structure of the receiver domain of RR02 from serotype 4 S. pneumoniae (as well as acetate- and phosphate-bound forms) at different pH levels. Two native structures at 2.3 Å, phased by single-wavelength anomalous diffraction (xenon SAD), and 1.85 Å and a third structure at pH 5.9 revealed the presence of a phosphate ion outside the active site. The fourth structure revealed the presence of an acetate molecule in the active site.

scholarly journals Biochemical Activities of the absA Two-Component System of Streptomyces coelicolor

2005 ◽  
Vol 187 (2) ◽  
pp. 687-696 ◽  
Author(s):  
Nancy L. Sheeler ◽  
Susan V. MacMillan ◽  
Justin R. Nodwell
Keyword(s):  
Transcriptional Repression ◽  
Kinase Activity ◽  
Response Regulator ◽  
Streptomyces Coelicolor ◽  
Point Mutations ◽  
Stable State ◽  
Sensor Kinase ◽  
Two Component System ◽  
Antibiotic Synthesis ◽  
Cognate Response Regulator

ABSTRACT The AbsA1 sensor kinase and its cognate response regulator AbsA2 are important regulators of antibiotic synthesis in Streptomyces coelicolor. While certain point mutations in absA1 reduce or eliminate the synthesis of several antibiotics, null mutations in these genes bring about enhanced antibiotic synthesis. We show here that AbsA1, which is unusual in sequence and structure, is both an AbsA2 kinase and an AbsA2∼P phosphatase. The half-life of AbsA2∼P in solution is 68.6 min, consistent with a role in maintaining a relatively stable state of transcriptional repression or activation. We find that mutations in the absA locus that enhance antibiotic synthesis impair AbsA2 kinase activity and that mutations that repress antibiotic synthesis impair AbsA2∼P phosphatase activity. These results support a model in which the phosphorylation state of AbsA2 is determined by the balance of the kinase and phosphatase activities of AbsA1 and where AbsA2∼P represses antibiotic biosynthetic genes either directly or indirectly.

scholarly journals Phosphorelay as the Sole Physiological Route of Signal Transmission by the Arc Two-Component System ofEscherichia coli

2000 ◽  
Vol 182 (13) ◽  
pp. 3858-3862 ◽  
Author(s):  
Ohsuk Kwon ◽  
Dimitris Georgellis ◽  
E. C. C. Lin
Keyword(s):  
Response Regulator ◽  
Signal Transmission ◽  
Phosphoryl Group ◽  
Sensor Kinase ◽  
Wild Type ◽  
Two Component System ◽  
Component System ◽  
Group Transfer ◽  
Two Component

ABSTRACT The Arc two-component system, comprising a tripartite sensor kinase (ArcB) and a response regulator (ArcA), modulates the expression of numerous genes involved in respiratory functions. In this study, the steps of phosphoryl group transfer from phosphorylated ArcB to ArcA were examined in vivo by using single copies of wild-type and mutantarcB alleles. The results indicate that the signal transmission occurs solely by His-Asp-His-Asp phosphorelay.

scholarly journals Crystal Structures of the Receiver Domain of the Response Regulator PhoP from Escherichia coli in the Absence and Presence of the Phosphoryl Analog Beryllofluoride

2007 ◽  
Vol 189 (16) ◽  
pp. 5987-5995 ◽  
Author(s):  
Priti Bachhawat ◽  
Ann M. Stock
Keyword(s):  
Escherichia Coli ◽  
Crystal Structures ◽  
Response Regulator ◽  
Active State ◽  
Two Component System ◽  
Component System ◽  
Regulatory Domains ◽  
Receiver Domain ◽  
Two Component ◽  
High Concentrations

ABSTRACT The response regulator PhoP is part of the PhoQ/PhoP two-component system involved in responses to depletion of extracellular Mg2+. Here, we report the crystal structures of the receiver domain of Escherichia coli PhoP determined in the absence and presence of the phosphoryl analog beryllofluoride. In the presence of beryllofluoride, the active receiver domain forms a twofold symmetric dimer similar to that seen in structures of other regulatory domains from the OmpR/PhoB family, providing further evidence that members of this family utilize a common mode of dimerization in the active state. In the absence of activating agents, the PhoP receiver domain crystallizes with a similar structure, consistent with the previous observation that high concentrations can promote an active state of PhoP independent of phosphorylation.

scholarly journals Phosphotransfer Reactions of the CbbRRS Three-Protein Two- Component System from Rhodopseudomonas palustris CGA010 Appear To Be Controlled by an Internal Molecular Switch on the Sensor Kinase

2006 ◽  
Vol 189 (2) ◽  
pp. 325-335 ◽  
Author(s):  
Simona Romagnoli ◽  
F. Robert Tabita
Keyword(s):  
Response Regulator ◽  
Rhodopseudomonas Palustris ◽  
Molecular Switch ◽  
Sensor Kinase ◽  
Two Component System ◽  
Conserved Residues ◽  
Component System ◽  
Receiver Domain ◽  
Two Component

ABSTRACT The CbbRRS system is an atypical three-protein two-component system that modulates the expression of the cbb I CO2 fixation operon of Rhodopseudomonas palustris, possibly in response to a redox signal. It consists of a membrane-bound hybrid sensor kinase, CbbSR, with a transmitter and receiver domain, and two response regulator proteins, CbbRR1 and CbbRR2. No detectable helix-turn-helix DNA binding domain is associated with either response regulator, but an HPt domain and a second receiver domain are predicted at the C-terminal region of CbbRR1 and CbbRR2, respectively. The abundance of conserved residues predicted to participate in a His-Asp phosphorelay raised the question of their de facto involvement. In this study, the role of the multiple receiver domains was elucidated in vitro by generating site-directed mutants of the putative conserved residues. Distinct phosphorylation patterns were obtained with two truncated versions of the hybrid sensor kinase, CbbSRT189 and CbbSRR96 (CbbSR beginning at residues T189 and R96, respectively). These constructs also exhibited substantially different affinities for ATP and phosphorylation stability, which was found to be dependent on a conserved Asp residue (Asp-696) within the kinase receiver domain. Asp-696 also played an important role in defining the specificity of phosphorylation for response regulators CbbRR1 or CbbRR2, and this residue appeared to act in conjunction with residues within the region from Arg-96 to Thr-189 at the N terminus of the sensor kinase. The net effect of concerted interactions at these distinct regions of CbbSR created an internal molecular switch that appears to coordinate a unique branched phosphorelay system.

scholarly journals Intra- and intermolecular domain interactions among novel two-component system proteins coded by Rv0600c, Rv0601c and Rv0602c of Mycobacterium tuberculosis

Microbiology ◽  
2009 ◽  
Vol 155 (3) ◽  
pp. 772-779 ◽  
Author(s):  
Rashmi Shrivastava ◽  
Ananta Kumar Ghosh ◽  
Amit Kumar Das
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Histidine Kinase ◽  
Response Regulator ◽  
Dominant Role ◽  
Phosphoryl Group ◽  
Two Component System ◽  
Component System ◽  
Two Component ◽  
Domain Interactions ◽  
Cognate Response Regulator

Two-component signal transduction pathways comprising a histidine kinase and its cognate response regulator play a dominant role in the adaptation of Mycobacterium tuberculosis to its host, and its virulence, pathogenicity and latency. Autophosphorylation occurs at a conserved histidine of the histidine kinase and subsequently the phosphoryl group is transferred to the conserved aspartate of its cognate response regulator. Among the twelve two-component systems of M. tuberculosis, Rv0600c (HK1), Rv0601c (HK2) and Rv0602c (TcrA) are annotated as a unique three-protein two-component system. HK1 contains an ATP-binding domain, and HK2, a novel Hpt mono-domain protein, contains the conserved phosphorylable histidine residue. HK1 and HK2 complement each other's functions. Interactions among different domains of the HK1, HK2 and TcrA proteins were studied using a yeast two-hybrid system. Self-interaction was observed for HK2 but not for HK1 or TcrA. HK2 was found to interact reasonably well with both HK1 and TcrA, but HK1 interacted weakly with TcrA. The conserved aspartate-containing receiver domain of TcrA interacted well with HK2 but not with HK1. These results suggest the existence of a novel signalling mechanism amongst HK1–HK2–TcrA, and a model for this mechanism is proposed.

scholarly journals Membrane Sensor Histidine Kinases: Insights from Structural, Ligand and Inhibitor Studies of Full-Length Proteins and Signalling Domains for Antibiotic Discovery

Molecules ◽  
2021 ◽  
Vol 26 (16) ◽  
pp. 5110
Author(s):  
Pikyee Ma ◽  
Mary K. Phillips-Jones
Keyword(s):  
Bacterial Infections ◽  
Environmental Changes ◽  
Structural Information ◽  
Response Regulator ◽  
Signal Transmission ◽  
Full Length ◽  
Bacterial Cells ◽  
Hydrophobic Membrane ◽  
Environmental Signals ◽  
Antibiotic Discovery

There is an urgent need to find new antibacterial agents to combat bacterial infections, including agents that inhibit novel, hitherto unexploited targets in bacterial cells. Amongst novel targets are two-component signal transduction systems (TCSs) which are the main mechanism by which bacteria sense and respond to environmental changes. TCSs typically comprise a membrane-embedded sensory protein (the sensor histidine kinase, SHK) and a partner response regulator protein. Amongst promising targets within SHKs are those involved in environmental signal detection (useful for targeting specific SHKs) and the common themes of signal transmission across the membrane and propagation to catalytic domains (for targeting multiple SHKs). However, the nature of environmental signals for the vast majority of SHKs is still lacking, and there is a paucity of structural information based on full-length membrane-bound SHKs with and without ligand. Reasons for this lack of knowledge lie in the technical challenges associated with investigations of these relatively hydrophobic membrane proteins and the inherent flexibility of these multidomain proteins that reduces the chances of successful crystallisation for structural determination by X-ray crystallography. However, in recent years there has been an explosion of information published on (a) methodology for producing active forms of full-length detergent-, liposome- and nanodisc-solubilised membrane SHKs and their use in structural studies and identification of signalling ligands and inhibitors; and (b) mechanisms of signal sensing and transduction across the membrane obtained using sensory and transmembrane domains in isolation, which reveal some commonalities as well as unique features. Here we review the most recent advances in these areas and highlight those of potential use in future strategies for antibiotic discovery. This Review is part of a Special Issue entitled “Interactions of Bacterial Molecules with Their Ligands and Other Chemical Agents” edited by Mary K. Phillips-Jones.

scholarly journals A unique genomic island-governed cannibalism in Bacillus enhanced biofilm formation through a novel regulation mechanism

2021 ◽  
Author(s):  
Rong Huang ◽  
Qing Li ◽  
Dandan Wang ◽  
Haichao Feng ◽  
Nan Zhang ◽  
...  
Keyword(s):  
Abc Transporter ◽  
Biofilm Formation ◽  
Response Regulator ◽  
Biological Significance ◽  
Signal Transduction Pathway ◽  
Phosphoryl Group ◽  
Regulation Mechanism ◽  
Two Component System ◽  
Differentiation Strategy ◽  
Bacterial Competition

Cannibalism is a differentiation strategy and social multicellular behavior in biofilms. The novel factors and mechanisms to trigger the bacterial cannibalism remain scarce. Here, we report a novel bacillunoic acids-mediated strategy for manipulating cannibalism in Bacillus velezensis SQR9 biofilm formation. A subfraction of cells differentiate into cannibals that secrete toxic bacillunoic acids to lyse a fraction of their sensitive siblings, and the released nutrients enhance biofilm formation. Meanwhile, the self-immunity of cannibal cells was induced by bacillunoic acids. A two-component system, the OmpS-OmpR signal-transduction pathway, controls the expression of the ABC transporter BnaAB for self-immunity. Specifically, bacillunoic acids activate the autophosphorylation of OmpS, a transmembrane histidine kinase, which then transfers a phosphoryl group to its response regulator OmpR. The phosphorylation of OmpR activates the transcription of the transporter gene bnaAB by binding its promoter. Thus, bacillunoic acids are pumped out of cells by the ABC transporter BnaAB. Moreover, we discovered that strain SQR9 could use the bacillunoic acids-mediated cannibalism to optimize its community to produce more bacillunoic acids for bacterial competition. This study revealed that bacillunoic acids play a previously undiscovered dual role in both cannibalism during biofilm formation and interspecies competition, which has an important biological significance.

scholarly journals Overexpression, purification, crystallization and preliminary X-ray crystallographic characterization of the receiver domain of the response regulator PhoP from Enterococcus faecalis ATCC 29212

2019 ◽  
Vol 62 (1) ◽  
Author(s):  
Yoon Chae Jeong ◽  
Ki Seog Lee
Keyword(s):  
Enterococcus Faecalis ◽  
Response Regulator ◽  
Critical Role ◽  
Transcriptional Response ◽  
Diffusion Method ◽  
Two Component System ◽  
Orthorhombic Space Group ◽  
Cell Parameters ◽  
Receiver Domain

Abstract Phosphate (Pho) regulon plays a critical role in bacterial phosphate homeostasis. It is regulated by two-component system (TCS) that comprises a sensor histidine kinase and transcriptional response regulator (RR). PhoP from Enterococcus faecalis (EfPhoP) belongs to the OmpR subfamily of RRs. It has not yet been structurally characterized because it is difficult to crystallize it to full-length form. In this study, a truncated form of EfPhoP containing the receiver domain (EfPhoP-RD) was constructed, purified to homogeneity and crystallized using the hanging-drop vapour-diffusion method. The crystal of EfPhoP-RD diffracted to 3.5 Å resolution and belonged to the orthorhombic space group C2221, with unit-cell parameters a = 118.74, b = 189.83, c = 189.88 Å. The asymmetric unit contains approximately 12 molecules, corresponding to a Matthews coefficient (Vm) of 2.50  Å3 Da−1 with a solvent content of 50.9%.

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