We demonstrated that it consists of two functional domains, domain A and domain B. NMR spectroscopic studies of these two domains revealed, for the first time, the structure of a histidine kinase. In this chapter, we describe functional and structural studies of EnvZc, which can be applied to characterize other histidine kinases. N2 - EnvZ is an osmosensing histidine kinase located in the inner membrane, and one of the most extensively studied Escherichia coli histidine kinases. AB - EnvZ is an osmosensing histidine kinase located in the inner membrane, and one of the most extensively studied Escherichia coli histidine kinases.
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Abstract EnvZ is an osmosensing histidine kinase located in the inner membrane, and one of the most extensively studied Escherichia coli histidine kinases. Fingerprint Histidine. Escherichia coli. Histidine Kinase. In addition, as the expression of many antibiotic resistance determinants is regulated by TCSs Poole, , an effective antimicrobial treatment could also be achieved by combining conventional and TCS-directed drugs.
Finally, a high degree of structural homology between the catalytic domains of the histidine kinase and the RR in bacteria suggest that multiple TCSs can be inhibited by a single compound, causing reduction in chromosomal resistance emergence Barrett and Isaacson, ; Poole, As TCS-encoding genes are found in all Gram-positive and Gram-negative bacterial genomes, a particular polypharmacological TCS inhibitor with the desired therapeutic effect may act comprehensively, in contrast to polypharmacy practice, if able to breach the various cell envelopes Anand and Chandra, In this review, we list several TCSs that are linked to bacterial virulence and antibiotic resistance, pointing out some of their main characteristics and addressing their possible therapeutic uses as drug targets.
Furthermore, the present review provides a compilation of TCS-inhibitors that have been identified by different assays of drug discovery. The basic biochemistry of the TCSs is well defined. A typical sensor histidine kinase comprises two domains: a variable N-terminal input domain and a conserved C-terminal domain that interacts with the RR. In turn, the RR contains one conserved N-terminal receiver domain and one variable C-terminal output domain Nixon et al. When stimulated, the sensor protein catalyzes autophosphorylation in a conserved histidine residue by using adenosine triphosphate ATP.
The high-energy phosphoryl group is then moved to a conserved aspartate residue of the RR, potentially changing its capability of targeting DNA sequences Bijlsma and Groisman, ; Cheung and Hendrickson, This signal transduction pathway has been reported to involve three phosphotransfer reactions and two phosphoprotein intermediates Stock et al. In recent years, since protein phosphorylation has been discovered to take place in bacterial nitrogen absorption and chemotaxis, a number of techniques have been developed to study Two-component signal transduction systems Scharf, PhoPQ is known to respond to a wide variety of environmental stress signals, including phosphate Ogura et al.
However, experimental verifications of stimuli that activate specific TCSs are reported in very few cases. When studying a TCS, a selection of candidate environmental conditions is usually made taking different physical and chemical parameters into consideration, such as changes in pH and osmolarity levels, oxygen pressure, temperature, exposure to ions, etc.
West and Stock, ; Varughese, It has also been reported that some TCSs present the ability to regulate gene clusters that contribute to cell growth, biofilm formation and virulence in pathogenic bacteria Eguchi and Utsumi, ; Mitrophanov and Groisman, ; Gotoh et al.
Nevertheless, in several cases, the role of TCSs in the pathogenicity of bacteria is not well understood and the attenuation of virulence is observed in TCS mutant strains without immediate knowledge of the exact mechanisms involved. Even though there is little explanation about the mechanisms involved in their attenuation, the TCS mutant strains present great potential to be used as live-attenuated vaccines against bacterial infections. For example, deletion of the genes encoding PhoP in Mycobacterium and Salmonella result in strains has been used as a vaccination strategy.
The phoP Salmonella mutants are attenuated and immunogenic for virulence in animal models VanCott et al. Hohmann et al. The association of TCSs with virulence has been studied in various pathogens, but few TCSs have been shown to be important for the coordination of expression of virulence factors.
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According to Sengupta et al. The arcA gene was mutated and experimental animal infections revealed that this gene positively controls the expression of toxT. The gene toxT encodes a transcriptional factor that increases the virulence of V. The large number of available bacterial genome sequence databases has made it possible to identify and predict interacting pairs of response regulatory kinases. Among the approaches are the use of tools of Next-generation sequencing NGS , molecular modeling, and bioinformatics.
For example, our group is working with the genomics of pathogenic bacteria. Corynebacterium pseudotuberculosis Cp is the etiological agent of Caseous Lymphadenitis CLA , an infectious disease that affects small ruminants worldwide Dorella et al. The genome sequence of this bacterium, recently completed, will aid in the identification of homologies and the prediction of novel genes that encode other virulence factors.
Genomic sequencing of C.
Circular representation of the genes encoding two-component systems TCSs in Corynebacterium pseudotuberculosis. A number of research groups have recently focused on TCSs to find potent inhibitors. In fact, many envelope transporter proteins acting as efflux pumps to promote the secretion of toxic compounds are controlled by TCSs. It has been reported that the efflux pumps regulated by TCSs in several important human pathogens, including Acinetobacter baumannii and Klebsiella pneumonia , provide multidrug resistance Howell et al.
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It has been reported that Staphylococcus aureus, Enterococcus faecalis and E. Targeting of this TCSs has led to the identification of several potent inhibitors that arrest the energy required for ATP synthesis in the cell Healy et al. Due to the negative effect, they exert over the mitochondrial respiration, these inhibitors could not be readily used as drugs. However, they may serve as structural templates for the discovery of novel selective inhibitors to target the VanSR TCS in adjunct therapy. As result, two anthraquinones produced by the plant species Rheum undulatum and R.
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Some predicted and experimentally validated compounds targeting bacterial TCSs are listed in Table 1. The principal approaches applied to the discovery of TCS inhibitors involve high throughput screening assays with purified TCSs and structure-based virtual screening SBVS using different types of compound libraries and rational drug design Qin et al. By using these techniques, a number of synthetic compounds belonging to several classes, such as benzoxazines, benzimidazoles Hilliard et al. TABLE 1. Two-component systems targeted by molecular compounds with antibacterial activity.
Presently, several factors contribute to the urgent need for innovative therapeutics: i the rise of anti-microbial resistance in pathogenic bacterial strains worldwide, ii prolonged therapy, iii co-morbidity with immunosuppressive diseases like HIV, pooled with a rapid rate of microbial evolution, iv the bacterial latency caused when bacteria enter and reside in a dormant state even for several decades and v a slower development of new antibiotics. In this regard, further basic research combined with modern drug discovery technologies may aid in the understanding of bacterial pathogenesis and cellular signaling pathways, leading to the development of novel therapeutic applications to exterminate bacterial infections.
Recognition of the relationship between TCSs and promotion of virulence in bacterial pathogens has directed the pharmaceutical industry to largely invest in the search of suitable inhibitors aiming signal transduction in bacteria. The catalytic and receiver domains of HKs and RRs are well studied and share a high degree of structural homology Barrett and Hoch, ; Domagala et al.
This knowledge suggests that a single drug compound capable of targeting any of these conserved domains could simultaneously inhibit multiple TCSs, boosting the chances to develop confrontation against the effects caused by the emergence of any molecular mutations affecting the affinity of the drug ligand for the target protein Okada et al.
In this regard, targeting multiple TCSs through their conserved domains may be more effective than targeting the varying sensor domain of each HK Gotoh et al. On the other hand, sequence conservation in kinase domains among bacteria and eukaryotes consists a possible disadvantage of targeting HKs. The ATP-binding pocket Bergerat fold present in bacterial HKs exhibits a high homology level with several human protein families and is also present in essential proteins found in a broad range of organisms, including the chaperone Hsp90 Bem et al.
A number of halogenated pyrrolo [2, 1-b] [1, 3] benzoxazines streptopyrroles isolated from fermentations of Streptomyces rimosus have been shown to possess antimicrobial activity against a series of bacteria and fungi Trew et al. The most promising streptopyrrole candidate presented inhibition against drug-resistant S. Stephenson and Hoch, However, similarities between the structures of streptopyrroles and salicylanilides suggest that the inhibition caused by these compounds may be a result of non-specific effects such as self-aggregation.
First, the SBVS analysis should be performed using compound databases comprehending a wide variety of potential inhibitors, including structures reported to present antibacterial activity Payne et al. Second, information about targeted TCSs should be obtained from clinically important pathogens to facilitate the identification of effective specific inhibitors.
Third, the modeling refinement of TCS protein structures may facilitate the search for compounds using the SBVS approach by allowing a more precise molecular docking analysis. The structural information obtained from these proteins can also help to identify the binding sites and chemical spaces that allow the interaction with candidate compounds, leading to further refinement in the prediction of novel inhibitors. In this regard, structural data of proteins that have been co-crystalized with ligand molecules are of special interest.
Finally, the generation of additional structure-activity relationship data will aid to minimize toxicity of the TCS inhibitors in eukaryotic cells and improve our understanding on the mechanisms of action of these compounds. The bacterial TCS consists of an important signaling mechanism for survival and establishment of pathogenic bacteria within the host. Along with the current need for new antimicrobial drugs, the regulatory nature of TCSs makes these systems outstanding targets for the development of alternative therapeutics against bacterial infections.
Their broad prevalence and the functional diversity of TCSs also support the need for testing several compounds against TCSs. In this regard, a better understanding of the interaction between a TCS and its targeting compound may aid in the development of an improved molecular structure presenting increased specificity for the correct ligand.
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Several works have reported natural and synthetic compounds presenting high affinity for TCSs and antimicrobial activity over pathogenic bacteria, but more studies are necessary to understand the exact mechanisms of action of these drugs. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
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Hye Suk Kong, Daniel P. Roberts, Cheryl D. Patterson, Sarah A. Kuehne, Stephan Heeb, Dilip K. Lakshman, and John Lydon. Characterization of each aefR and mexT mutant in Pseudomonas syringae pv. Microarray analysis and phenotypic response of Pseudomonas aeruginosa PAO1 under hyperbaric oxyhelium conditions. GacA directly regulates expression of several virulence genes in Pseudomonas syringae pv. Stringent response mutants of Pseudomonas chlororaphis PA23 exhibit enhanced antifungal activity against Sclerotinia sclerotiorum in vitro. P2TF: a comprehensive resource for analysis of prokaryotic transcription factors.
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Molecular communication in the rhizosphere. Temperature-responsive sensing regulates biocontrol factor expression in Pseudomonas fluorescens CHA0. Ultramicrocells form by reductive division in macroscopic Pseudomonas aerial structures. Prokaryote-eukaryote interactions identified by using Caenorhabditis elegans.
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Phytase activity and its regulation in a rhizospheric strain of Serratia plymuthica. Central role of quorum sensing in regulating the production of pathogenicity factors in Pseudomonas aeruginosa. Reedy, Amy O. Charkowski, and Ching-Hong Yang. The post-transcriptional regulator CsrA plays a central role in the adaptation of bacterial pathogens to different stages of infection in animal hosts. Metabolites produced by Pseudomonas sp. Effect of mutations in global regulator genes grrS and rpoS on synthesis of phosphatases in Serratia plymuthica. Dialogues of root-colonizing biocontrol pseudomonads.
Interactions between plants and beneficial Pseudomonas spp.
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