Blue Light Regulated Two-Component Systems: Enzymatic and Functional Analyses of Light-Oxygen-Voltage (LOV)-Histidine Kinases and Downstream Response Regulators

Two-component systems (TCS) are protein machineries that enable cells to respond to input signals. Histidine kinases (HK) are the sensory component, transferring information toward downstream response regulators (RR). HKs transfer phosphoryl groups to their specific RRs, but also dephosphorylate them, overall ensuring proper signaling. The mechanisms by which HKs discriminate between such disparate directions, are yet unknown. We now disclose crystal structures of the HK:RR complex DesK:DesR from Bacillus subtilis, comprising snapshots of the phosphotransfer and the dephosphorylation reactions. The HK dictates the reactional outcome through conformational rearrangements that include the reactive histidine. The phosphotransfer center is asymmetric, poised for dissociative nucleophilic substitution. The structural bases of HK phosphatase/phosphotransferase control are uncovered, and the unexpected discovery of a dissociative reactional center, sheds light on the evolution of TCS phosphotransfer reversibility. Our findings should be applicable to a broad range of signaling systems and instrumental in synthetic TCS rewiring.

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Specificity Residues Determine Binding Affinity for Two-Component Signal Transduction Systems

mBio ◽

10.1128/mbio.00420-13 ◽

2013 ◽

Vol 4 (6) ◽

Cited By ~ 32

Author(s):

Jonathan W. Willett ◽

Nitija Tiwari ◽

Susanne Müller ◽

Katherine R. Hummels ◽

Jon C. D. Houtman ◽

...

Keyword(s):

Binding Affinity ◽

Myxococcus Xanthus ◽

Titration Calorimetry ◽

Signaling Proteins ◽

Response Regulators ◽

Histidine Kinases ◽

Content Type ◽

Component Systems ◽

Two Component ◽

Two Component Systems

ABSTRACTTwo-component systems (TCS) comprise histidine kinases and their cognate response regulators and allow bacteria to sense and respond to a wide variety of signals. Histidine kinases (HKs) phosphorylate and dephosphorylate their cognate response regulators (RRs) in response to stimuli. In general, these reactions appear to be highly specific and require an appropriate association between the HK and RR proteins. TheMyxococcus xanthusgenome encodes one of the largest repertoires of signaling proteins in bacteria (685 open reading frames [ORFs]), including at least 127 HKs and at least 143 RRs. Of these, 27 arebona fideNtrC-family response regulators, 21 of which are encoded adjacent to their predicted cognate kinases. Using system-wide profiling methods, we determined that the HK-NtrC RR pairs display a kinetic preference during both phosphotransfer and phosphatase functions, thereby defining cognate signaling systems inM. xanthus. Isothermal titration calorimetry measurements indicated that cognate HK-RR pairs interact with dissociation constants (Kd) of approximately 1 µM, while noncognate pairs had no measurable binding. Lastly, a chimera generated between the histidine kinase, CrdS, and HK1190 revealed that residues conferring phosphotransfer and phosphatase specificity dictate binding affinity, thereby establishing discrete protein-protein interactions which prevent cross talk. The data indicate that binding affinity is a critical parameter governing system-wide signaling fidelity for bacterial signal transduction proteins.IMPORTANCEUsingin vitrophosphotransfer and phosphatase profiling assays and isothermal titration calorimetry, we have taken a system-wide approach to demonstrate specificity for a family of two-component signaling proteins inMyxococcus xanthus. Our results demonstrate that previously identified specificity residues dictate binding affinity and that phosphatase specificity follows phosphotransfer specificity for cognate HK-RR pairs. The data indicate that preferential binding affinity is the basis for signaling fidelity in bacterial two-component systems.

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Unequal twins: Unraveling the reaction mechanism of dimeric histidine kinases

Journal of Biological Chemistry ◽

10.1074/jbc.h120.014170 ◽

2020 ◽

Vol 295 (23) ◽

pp. 8118-8119

Author(s):

Wolfgang Gärtner

Keyword(s):

Basic Function ◽

Response Regulators ◽

Histidine Kinases ◽

Open Questions ◽

Component Systems ◽

Two Component ◽

New Strategy ◽

Two Component Systems ◽

Future Work ◽

Partner Proteins

Histidine kinases (HKs), together with their partner proteins, the response regulators (RRs), form the ubiquitous two-component systems that are global players in control and adjustment of microbial lifestyle. Although their basic function (i.e. the transfer of a phosphate group from the HK to its RR partner) is simple to articulate, deciphering the molecular details of this process has proven anything but simple, especially when quantitative aspects come into play. Bouillet et al. report a series of elegant and sophisticated experiments to quantitatively understand HK functions, clearing up several open questions and providing a new strategy for future work in the field.

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Two-Component Sensing and Regulation: How Do Histidine Kinases Talk with Response Regulators at the Molecular Level?

Annual Review of Microbiology ◽

10.1146/annurev-micro-091018-054627 ◽

2019 ◽

Vol 73 (1) ◽

pp. 507-528 ◽

Cited By ~ 17

Author(s):

Alejandro Buschiazzo ◽

Felipe Trajtenberg

Keyword(s):

Molecular Level ◽

Response Regulator ◽

Response Regulators ◽

Histidine Kinases ◽

Universal Principles ◽

Component Systems ◽

Two Component ◽

Two Component Systems ◽

Internal Information ◽

Living Organisms

Perceiving environmental and internal information and reacting in adaptive ways are essential attributes of living organisms. Two-component systems are relevant protein machineries from prokaryotes and lower eukaryotes that enable cells to sense and process signals. Implicating sensory histidine kinases and response regulator proteins, both components take advantage of protein phosphorylation and flexibility to switch conformations in a signal-dependent way. Dozens of two-component systems act simultaneously in any given cell, challenging our understanding about the means that ensure proper connectivity. This review dives into the molecular level, attempting to summarize an emerging picture of how histidine kinases and cognate response regulators achieve required efficiency, specificity, and directionality of signaling pathways, properties that rely on protein:protein interactions. α helices that carry information through long distances, the fine combination of loose and specific kinase/regulator interactions, and malleable reaction centers built when the two components meet emerge as relevant universal principles.

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Conformational dynamics of the essential sensor histidine kinase WalK

Acta Crystallographica Section D Structural Biology ◽

10.1107/s2059798317013043 ◽

2017 ◽

Vol 73 (10) ◽

pp. 793-803 ◽

Cited By ~ 12

Author(s):

Yongfei Cai ◽

Mingyang Su ◽

Ashfaq Ahmad ◽

Xiaojie Hu ◽

Jiayan Sang ◽

...

Keyword(s):

Dynamic Properties ◽

Conformational Dynamics ◽

Bound State ◽

Response Regulators ◽

Histidine Kinases ◽

Sequence Identity ◽

Component Systems ◽

Bacterial Signal Transduction ◽

Two Component Systems ◽

Intrinsic Dynamic

Two-component systems (TCSs) are key elements in bacterial signal transduction in response to environmental stresses. TCSs generally consist of sensor histidine kinases (SKs) and their cognate response regulators (RRs). Many SKs exhibit autokinase, phosphoryltransferase and phosphatase activities, which regulate RR activity through a phosphorylation and dephosphorylation cycle. However, how SKs perform different enzymatic activities is poorly understood. Here, several crystal structures of the minimal catalytic region of WalK, an essential SK fromLactobacillus plantarumthat shares 60% sequence identity with its homologue VicK fromStreptococcus mutans, are presented. WalK adopts an asymmetrical closed structure in the presence of ATP or ADP, in which one of the CA domains is positioned close to the DHp domain, thus leading both the β- and γ-phosphates of ATP/ADP to form hydrogen bonds to the ∊- but not the δ-nitrogen of the phosphorylatable histidine in the DHp domain. In addition, the DHp domain in the ATP/ADP-bound state has a 25.7° asymmetrical helical bending coordinated with the repositioning of the CA domain; these processes are mutually exclusive and alternate in response to helicity changes that are possibly regulated by upstream signals. In the absence of ATP or ADP, however, WalK adopts a completely symmetric open structure with its DHp domain centred between two outward-reaching CA domains. In summary, these structures of WalK reveal the intrinsic dynamic properties of an SK structure as a molecular basis for multifunctionality.

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Molecular Characterization of Two-Component Systems of Helicobacter pylori

Journal of Bacteriology ◽

10.1128/jb.182.8.2068-2076.2000 ◽

2000 ◽

Vol 182 (8) ◽

pp. 2068-2076 ◽

Cited By ~ 126

Author(s):

Dagmar Beier ◽

Rainer Frank

Keyword(s):

Helicobacter Pylori ◽

Histidine Kinase ◽

Response Regulator ◽

Response Regulators ◽

Reading Frame ◽

Component Systems ◽

Two Component ◽

Two Component Systems

ABSTRACT Two-component systems are frequently involved in the adaptation of bacteria to changing environmental conditions at the level of transcriptional regulation. Here we report the characterization of members of the two-component systems of the gastric pathogenHelicobacter pylori deduced from the genome sequence of strain 26695. We demonstrate that the response regulators HP166, HP1043, and HP1021 have essential functions, as disruption of the corresponding genes is lethal for the bacteria, irrespective of the fact that HP1043 and HP1021 have nonconserved substitutions in crucial amino acids of their receiver domains. An analysis of the in vitro phosphorylation properties of the two-component proteins demonstrates that HP244-HP703 and HP165-HP166 are cognate histidine kinase-response regulator pairs. Furthermore, we provide evidence that the variability of the histidine kinase HP165 caused by a poly(C) tract of variable length close to the 3′ end of open reading frame 165/164 does not interfere with the kinase activity of the transmitter domain of HP165.

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Two-Component Systems of S. aureus: Signaling and Sensing Mechanisms

Genes ◽

10.3390/genes13010034 ◽

2021 ◽

Vol 13 (1) ◽

pp. 34

Author(s):

Lisa Bleul ◽

Patrice Francois ◽

Christiane Wolz

Keyword(s):

Transcriptional Activation ◽

Drug Targets ◽

Bacterial Survival ◽

Histidine Kinases ◽

Signal Perception ◽

Signaling Mechanisms ◽

Component Systems ◽

Two Component ◽

Two Component Systems ◽

Sense And Respond

Staphylococcus aureus encodes 16 two-component systems (TCSs) that enable the bacteria to sense and respond to changing environmental conditions. Considering the function of these TCSs in bacterial survival and their potential role as drug targets, it is important to understand the exact mechanisms underlying signal perception. The differences between the sensing of appropriate signals and the transcriptional activation of the TCS system are often not well described, and the signaling mechanisms are only partially understood. Here, we review present insights into which signals are sensed by histidine kinases in S. aureus to promote appropriate gene expression in response to diverse environmental challenges.

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Unique Patterns and Biogeochemical Relevance of Two-Component Sensing in Marine Bacteria

mSystems ◽

10.1128/msystems.00317-18 ◽

2019 ◽

Vol 4 (1) ◽

Cited By ~ 7

Author(s):

Noelle A. Held ◽

Matthew R. McIlvin ◽

Dawn M. Moran ◽

Michael T. Laub ◽

Mak A. Saito

Keyword(s):

Marine Environment ◽

Marine Bacteria ◽

Response Regulator ◽

Bacterial Species ◽

South Pacific ◽

Response Regulators ◽

Orphan Genes ◽

Component Systems ◽

Two Component ◽

Two Component Systems

ABSTRACTTwo-component sensory (TCS) systems link microbial physiology to the environment and thus may play key roles in biogeochemical cycles. In this study, we surveyed the TCS systems of 328 diverse marine bacterial species. We identified lifestyle traits such as copiotrophy and diazotrophy that are associated with larger numbers of TCS system genes within the genome. We compared marine bacterial species with 1,152 reference bacterial species from a variety of habitats and found evidence of extra response regulators in marine genomes. Examining the location of TCS genes along the circular bacterial genome, we also found that marine bacteria have a large number of “orphan” genes, as well as many hybrid histidine kinases. The prevalence of “extra” response regulators, orphan genes, and hybrid TCS systems suggests that marine bacteria break with traditional understanding of how TCS systems operate. These trends suggest prevalent regulatory networking, which may allow coordinated physiological responses to multiple environmental signals and may represent a specific adaptation to the marine environment. We examine phylogenetic and lifestyle traits that influence the number and structure of two-component systems in the genome, finding, for example, that a lack of two-component systems is a hallmark of oligotrophy. Finally, in an effort to demonstrate the importance of TCS systems to marine biogeochemistry, we examined the distribution ofProchlorococcus/Synechococcusresponse regulator PMT9312_0717 in metaproteomes of the tropical South Pacific. We found that this protein’s abundance is related to phosphate concentrations, consistent with a putative role in phosphate regulation.IMPORTANCEMarine microbes must manage variation in their chemical, physical, and biological surroundings. Because they directly link bacterial physiology to environmental changes, TCS systems are crucial to the bacterial cell. This study surveyed TCS systems in a large number of marine bacteria and identified key phylogenetic and lifestyle patterns in environmental sensing. We found evidence that, in comparison with bacteria as a whole, marine organisms have irregular TCS system constructs which might represent an adaptation specific to the marine environment. Additionally, we demonstrate the biogeochemical relevance of TCS systems by correlating the presence of the PMT9312_0717 response regulator protein to phosphate concentrations in the South Pacific. We highlight that despite their potential ecological and biogeochemical relevance, TCS systems have been understudied in the marine ecosystem. This report expands our understanding of the breadth of bacterial TCS systems and how marine bacteria have adapted to survive in their unique environment.

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