scholarly journals IN SILICO ANALYSIS AND FUNCTIONAL CHARACTERIZATION OF FHUB, A COMPONENT OF ERWINIA MALLOTIVORA FERRIC HYDROXAMATE UPTAKE SYSTEM

2020 ◽  
Vol 82 (3) ◽  
Author(s):  
Nor Mustaiqazah Juri ◽  
Aimera Farhana Samsuddin ◽  
Abdul Munir Abd Murad ◽  
Amin Asyraf Tamizi ◽  
Mohd Azhar Hassan ◽  
...  
Keyword(s):  
In Silico ◽  
Pathogenic Bacteria ◽  
Iron Acquisition ◽  
Functional Characterization ◽  
Membrane Topology ◽  
Transport Systems ◽  
Critical Element ◽  
Uptake System ◽  
Dieback Disease ◽  
Ferric Hydroxamate

Iron is a critical element for bacterial growth as most pathogenic bacteria relies on their host for iron supply. However, iron sources are bounded to the host iron binding protein and specific iron acquisition mechanism is required to chelate and transport the iron to the bacteria. Ferric hydroxamate uptake system or fhu is one of the transport systems that import iron in the form of ferric hydroxamate/ ferrichrome from the extracellular environment into the bacterial cytosol. In this present study, a detailed in silico structural analysis was conducted on an important component of fhu transport member from Erwinia mallotivora named as fhuB. This provide us the structural properties of the protein which includes the domain and 3D model, phylogenetic analysis and the membrane topology. For functional analysis, a knockout mutant of fhuB gene strain was generated to evaluate the effect of silencing this gene during E. mallotivora infection in papaya. When compared to the wild E. mallotivora strain, fhuB mutant strain of E. mallotivora loss its virulence in causing dieback disease symptom in papaya. The result of this study has revealed the significant role of iron acquisition and metabolism during E. mallotivora pathogenesis. This highlights fhuB role and importance as the target gene; to inhibit iron uptake in E. mallotivora for future study and as a part of future consideration for dieback disease management strategy in papaya.

scholarly journals Identification and Regulation of Genes for Cobalamin Transport in the Cyanobacterium Synechococcus sp. Strain PCC 7002

2016 ◽  
Vol 198 (19) ◽  
pp. 2753-2761 ◽  
Author(s):  
Adam A. Pérez ◽  
Dmitry A. Rodionov ◽  
Donald A. Bryant
Keyword(s):  
Active Transport ◽  
Transport System ◽  
In Silico ◽  
Fluorescent Protein ◽  
Yellow Fluorescent Protein ◽  
Transport Systems ◽  
Uptake System ◽  
Reporter System ◽  
Content Type ◽  
Active Transport System

ABSTRACTThe cyanobacteriumSynechococcussp. strain PCC 7002 is a cobalamin auxotroph and utilizes this coenzyme solely for the synthesis ofl-methionine by methionine synthase (MetH).Synechococcussp. strain PCC 7002 is unable to synthesize cobalaminde novo, and because of the large size of this tetrapyrrole, an active-transport system must exist for cobalamin uptake. Surprisingly, no cobalamin transport system was identified in the initial annotation of the genome of this organism. With more sophisticatedin silicoprediction tools, abtuB-cpdA-btuC-btuFoperon encoding components putatively required for a B12uptake (btu) system was identified. The expression of these genes was predicted to be controlled by a cobalamin riboswitch. Global transcriptional profiling by high-throughput RNA sequencing of a cobalamin-independent form ofSynechococcussp. strain PCC 7002 grown in the absence or presence of cobalamin confirmed regulation of thebtuoperon by cobalamin. Pérez et al. (A. A. Pérez, Z. Liu, D. A. Rodionov, Z. Li, and D. A. Bryant, J Bacteriol 198:2743–2752, 2016,http://dx.doi.org/10.1128/JB.00475-16) developed a cobalamin-dependent yellow fluorescent protein reporter system in aSynechococcussp. strain PCC 7002 variant that had been genetically modified to allow cobalamin-independent growth. This reporter system was exploited to validate components of thebtuuptake system by assessing the ability of targeted mutants to transport cobalamin. ThebtuBpromoter and a variant counterpart mutated in an essential element of the predicted cobalamin riboswitch were fused to ayfpreporter. The combined data indicate that thebtuB-cpdA-btuF-btuCoperon in this cyanobacterium is transcriptionally regulated by a cobalamin riboswitch.IMPORTANCEWith a cobalamin-regulated reporter system for expression of yellow fluorescent protein, genes previously misidentified as encoding subunits of a siderophore transporter were shown to encode components of cobalamin uptake in the cyanobacteriumSynechococcussp. strain PCC 7002. This study demonstrates the importance of experimental validation ofin silicopredictions and provides a general scheme forin vivoverification of similar cobalamin transport systems. A putative cobalamin riboswitch was identified inSynechococcussp. strain PCC 7002. This riboswitch acts as a potential transcriptional attenuator of thebtuoperon that encodes the components of the cobalamin active-transport system.

scholarly journals Bacterial iron acquisition mediated by outer membrane translocation and cleavage of a host protein

2018 ◽  
Vol 115 (26) ◽  
pp. 6840-6845 ◽  
Author(s):  
Khedidja Mosbahi ◽  
Marta Wojnowska ◽  
Amaya Albalat ◽  
Daniel Walker
Keyword(s):  
Cell Surface ◽  
Outer Membrane ◽  
Pathogenic Bacteria ◽  
Protein Import ◽  
Binding Protein ◽  
Iron Acquisition ◽  
Host Protein ◽  
Transport Systems ◽  
Growth Enhancement ◽  
Periplasmic Binding Protein

Iron is an essential micronutrient for most bacteria and is obtained from iron-chelating siderophores or directly from iron-containing host proteins. For Gram-negative bacteria, classical iron transport systems consist of an outer membrane receptor, a periplasmic binding protein, and an inner membrane ABC transporter, which work in concert to deliver iron from the cell surface to the cytoplasm. We recently showed thatPectobacteriumspp. are able to acquire iron from ferredoxin, a small and stable 2Fe-2S iron sulfur cluster containing protein and identified the ferredoxin receptor, FusA, a TonB-dependent receptor that binds ferredoxin on the cell surface. The genetic context offusAsuggests an atypical iron acquisition system, lacking a periplasmic binding protein, although the mechanism through which iron is extracted from the captured ferredoxin has remained unknown. Here we show that FusC, an M16 family protease, displays a highly targeted proteolytic activity against plant ferredoxin, and that growth enhancement ofPectobacteriumdue to iron acquisition from ferredoxin is FusC-dependent. The periplasmic location of FusC indicates a mechanism in which ferredoxin is imported into the periplasm via FusA before cleavage by FusC, as confirmed by the uptake and accumulation of ferredoxin in the periplasm in a strain lackingfusC. The existence of homologous uptake systems in a range of pathogenic bacteria suggests that protein uptake for nutrient acquisition may be widespread in bacteria and shows that, similar to their endosymbiotic descendants mitochondria and chloroplasts, bacteria produce dedicated protein import systems.

Acetate Kinase (AcK) is Essential for Microbial Growth and Betel-derived Compounds Potentially Target AcK, PhoP and MDR Proteins in M. tuberculosis, V. cholerae and Pathogenic E. coli: An in silico and in vitro Study

2019 ◽  
Vol 18 (31) ◽  
pp. 2731-2740 ◽  
Author(s):  
Sandeep Tiwari ◽  
Debmalya Barh ◽  
M. Imchen ◽  
Eswar Rao ◽  
Ranjith K. Kumavath ◽  
...  
Keyword(s):  
Broad Spectrum ◽  
In Silico ◽  
Pathogenic Bacteria ◽  
In Vitro Study ◽  
Docking Studies ◽  
Acetate Kinase ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Novel Target

Background: Mycobacterium tuberculosis, Vibrio cholerae, and pathogenic Escherichia coli are global concerns for public health. The emergence of multi-drug resistant (MDR) strains of these pathogens is creating additional challenges in controlling infections caused by these deadly bacteria. Recently, we reported that Acetate kinase (AcK) could be a broad-spectrum novel target in several bacteria including these pathogens. Methods: Here, using in silico and in vitro approaches we show that (i) AcK is an essential protein in pathogenic bacteria; (ii) natural compounds Chlorogenic acid and Pinoresinol from Piper betel and Piperidine derivative compound 6-oxopiperidine-3-carboxylic acid inhibit the growth of pathogenic E. coli and M. tuberculosis by targeting AcK with equal or higher efficacy than the currently used antibiotics; (iii) molecular modeling and docking studies show interactions between inhibitors and AcK that correlate with the experimental results; (iv) these compounds are highly effective even on MDR strains of these pathogens; (v) further, the compounds may also target bacterial two-component system proteins that help bacteria in expressing the genes related to drug resistance and virulence; and (vi) finally, all the tested compounds are predicted to have drug-like properties. Results and Conclusion: Suggesting that, these Piper betel derived compounds may be further tested for developing a novel class of broad-spectrum drugs against various common and MDR pathogens.

scholarly journals In Silico Analyses of Rice Thionin Genes and the Antimicrobial Activity of OsTHION15 Against Phytopathogens

2019 ◽  
Vol 109 (1) ◽  
pp. 27-35
Author(s):  
Krissana Boonpa ◽  
Suparuk Tantong ◽  
Kamonwan Weerawanich ◽  
Pawinee Panpetch ◽  
Onanong Pringsulaka ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Stress Responses ◽  
In Silico ◽  
Pathogenic Bacteria ◽  
Hyphal Growth ◽  
In Planta ◽  
Recombinant Peptide ◽  
Plant Disease Control ◽  
Helminthosporium Oryzae ◽  
In Silico Analyses

Thionins are a family of antimicrobial peptides. We performed in silico expression analyses of the 44 rice (Oryza sativa) thionins (OsTHIONs). Modulated expression levels of OsTHIONs under different treatments suggest their involvement in many processes, including biotic, abiotic, and nutritional stress responses, and in hormone signaling. OsTHION15 (LOC_Os06g32600) was selected for further characterization based on several in silico analyses. OsTHION15 in O. sativa subsp. indica ‘KDML 105’ was expressed in all of the tissues and organs examined, including germinating seed, leaves, and roots of seedlings and mature plants, and inflorescences. To investigate the antimicrobial activity of OsTHION15, we produced a recombinant peptide in Escherichia coli Rosetta-gami (DE3). The recombinant OsTHION15 exhibited inhibitory activities toward rice-pathogenic bacteria such as Xanthomonas oryzae pv. oryzae and Pectobacterium carotovorum pv. atroseptica, with minimum inhibitory concentrations of 112.6 and 14.1 µg ml−1, respectively. A significant hyphal growth inhibition was also observed toward Fusarium oxysporum f. sp. cubense and Helminthosporium oryzae. In addition, we demonstrated the in planta antibacterial activity of this peptide in Nicotiana benthamiana against X. campestris pv. glycines. These activities suggest the possible application of OsTHION15 in plant disease control.

scholarly journals Metabolic Modeling of Pectobacterium parmentieri SCC3193 Provides Insights into Metabolic Pathways of Plant Pathogenic Bacteria

2019 ◽  
Vol 7 (4) ◽  
pp. 101 ◽  
Author(s):  
Sabina Zoledowska ◽  
Luana Presta ◽  
Marco Fondi ◽  
Francesca Decorosi ◽  
Luciana Giovannetti ◽  
...  
Keyword(s):  
In Silico ◽  
Pathogenic Bacteria ◽  
Metabolic Modeling ◽  
Metabolic Model ◽  
Metabolic Adaptation ◽  
Ecological Niches ◽  
Plant Colonization ◽  
Phenotypic Data ◽  
Plant Pathogenic Bacteria ◽  
Metabolic Versatility

Understanding plant–microbe interactions is crucial for improving plants’ productivity and protection. Constraint-based metabolic modeling is one of the possible ways to investigate the bacterial adaptation to different ecological niches and may give insights into the metabolic versatility of plant pathogenic bacteria. We reconstructed a raw metabolic model of the emerging plant pathogenic bacterium Pectobacterium parmentieri SCC3193 with the use of KBase. The model was curated by using inParanoind and phenotypic data generated with the use of the OmniLog system. Metabolic modeling was performed through COBRApy Toolbox v. 0.10.1. The curated metabolic model of P. parmentieri SCC3193 is highly reliable, as in silico obtained results overlapped up to 91% with experimental data on carbon utilization phenotypes. By mean of flux balance analysis (FBA), we predicted the metabolic adaptation of P. parmentieri SCC3193 to two different ecological niches, relevant for the persistence and plant colonization by this bacterium: soil and the rhizosphere. We performed in silico gene deletions to predict the set of essential core genes for this bacterium to grow in such environments. We anticipate that our metabolic model will be a valuable element for defining a set of metabolic targets to control infection and spreading of this plant pathogen.

scholarly journals Trk2 Potassium Transport System in Streptococcus mutans and Its Role in Potassium Homeostasis, Biofilm Formation, and Stress Tolerance

2016 ◽  
Vol 198 (7) ◽  
pp. 1087-1100 ◽  
Author(s):  
Gursonika Binepal ◽  
Kamal Gill ◽  
Paula Crowley ◽  
Martha Cordova ◽  
L. Jeannine Brady ◽  
...  
Keyword(s):  
Stress Tolerance ◽  
Streptococcus Mutans ◽  
Transport System ◽  
Biofilm Formation ◽  
Cellular Response ◽  
Pathogenic Bacteria ◽  
Transport Systems ◽  
Content Type ◽  
Growth And Survival ◽  
Dental Biofilm

ABSTRACTPotassium (K+) is the most abundant cation in the fluids of dental biofilm. The biochemical and biophysical functions of K+and a variety of K+transport systems have been studied for most pathogenic bacteria but not for oral pathogens. In this study, we establish the modes of K+acquisition inStreptococcus mutansand the importance of K+homeostasis for its virulence attributes. TheS. mutansgenome harbors four putative K+transport systems that included two Trk-like transporters (designated Trk1 and Trk2), one glutamate/K+cotransporter (GlnQHMP), and a channel-like K+transport system (Kch). Mutants lacking Trk2 had significantly impaired growth, acidogenicity, aciduricity, and biofilm formation. [K+] less than 5 mM eliminated biofilm formation inS. mutans. The functionality of the Trk2 system was confirmed by complementing anEscherichia coliTK2420 mutant strain, which resulted in significant K+accumulation, improved growth, and survival under stress. Taken together, these results suggest that Trk2 is the main facet of the K+-dependent cellular response ofS. mutansto environment stresses.IMPORTANCEBiofilm formation and stress tolerance are important virulence properties of caries-causingStreptococcus mutans. To limit these properties of this bacterium, it is imperative to understand its survival mechanisms. Potassium is the most abundant cation in dental plaque, the natural environment ofS. mutans. K+is known to function in stress tolerance, and bacteria have specialized mechanisms for its uptake. However, there are no reports to identify or characterize specific K+transporters inS. mutans. We identified the most important system for K+homeostasis and its role in the biofilm formation, stress tolerance, and growth. We also show the requirement of environmental K+for the activity of biofilm-forming enzymes, which explains why such high levels of K+would favor biofilm formation.

scholarly journals Staphylococcus aureus Serves as an Iron Source for Pseudomonas aeruginosa during In Vivo Coculture

2005 ◽  
Vol 187 (2) ◽  
pp. 554-566 ◽  
Author(s):  
Lauren M. Mashburn ◽  
Amy M. Jett ◽  
Darrin R. Akins ◽  
Marvin Whiteley
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Pathogenic Bacteria ◽  
Iron Acquisition ◽  
Low Oxygen ◽  
Dialysis Membrane ◽  
Opportunistic Human Pathogen ◽  
The Impact

ABSTRACT Pseudomonas aeruginosa is a gram-negative opportunistic human pathogen often infecting the lungs of individuals with the heritable disease cystic fibrosis and the peritoneum of individuals undergoing continuous ambulatory peritoneal dialysis. Often these infections are not caused by colonization with P. aeruginosa alone but instead by a consortium of pathogenic bacteria. Little is known about growth and persistence of P. aeruginosa in vivo, and less is known about the impact of coinfecting bacteria on P. aeruginosa pathogenesis and physiology. In this study, a rat dialysis membrane peritoneal model was used to evaluate the in vivo transcriptome of P. aeruginosa in monoculture and in coculture with Staphylococcus aureus. Monoculture results indicate that approximately 5% of all P. aeruginosa genes are differentially regulated during growth in vivo compared to in vitro controls. Included in this analysis are genes important for iron acquisition and growth in low-oxygen environments. The presence of S. aureus caused decreased transcription of P. aeruginosa iron-regulated genes during in vivo coculture, indicating that the presence of S. aureus increases usable iron for P. aeruginosa in this environment. We propose a model where P. aeruginosa lyses S. aureus and uses released iron for growth in low-iron environments.

scholarly journals Neutrophil Gelatinase-Associated Lipocalin Expresses Antimicrobial Activity by Interfering with l-Norepinephrine-Mediated Bacterial Iron Acquisition

2010 ◽  
Vol 54 (4) ◽  
pp. 1580-1589 ◽  
Author(s):  
Marcus Miethke ◽  
Arne Skerra
Keyword(s):  
Iron Acquisition ◽  
Primary Source ◽  
Iron Complexes ◽  
Iron Complex ◽  
Immune Defense ◽  
Uptake System ◽  
Iron Chelate ◽  
Neutrophil Gelatinase Associated Lipocalin ◽  
Bacterial Uptake ◽  
Neutrophil Gelatinase

ABSTRACT l-norepinephrine (NE) is a neuroendocrine catecholamine that supports bacterial growth by mobilizing iron from a primary source such as holotransferrin to increase its bioavailability for cellular uptake. Iron complexes of NE resemble those of bacterial siderophores that are scavenged by human neutrophil gelatinase-associated lipocalin (NGAL) as part of the innate immune defense. Here, we show that NGAL binds iron-complexed NE, indicating physiological relevance for both bacterial and human iron metabolism. The fluorescence titration of purified recombinant NGAL with the FeIII·(NE)3 iron complex revealed high affinity for this ligand, with a K D of 50.6 nM. In contrast, the binding protein FeuA of Bacillus subtilis, which is involved in the bacterial uptake of triscatecholate iron complexes, has a K D for FeIII·(NE)3 of 1.6 μM, indicating that NGAL is an efficient competitor. Furthermore, NGAL was shown to inhibit the NE-mediated growth of both E. coli and B. subtilis strains that either are capable or incapable of producing their native siderophores enterobactin and bacillibactin, respectively. These experiments suggest that iron-complexed NE directly serves as an iron source for bacterial uptake systems, and that NGAL can function as an antagonist of this iron acquisition process. Interestingly, a functional FeuABC uptake system was shown to be necessary for NE-mediated growth stimulation as well as its NGAL-dependent inhibition. This study demonstrates for the first time that human NGAL not only neutralizes pathogen-derived virulence factors but also can effectively scavenge an iron-chelate complex abundant in the host.

G-quadruplex motifs are functionally conserved in cis-regulatory regions of pathogenic bacteria: An in-silico evaluation

Biochimie ◽  
2021 ◽  
Vol 184 ◽  
pp. 40-51
Author(s):  
Upalabdha Dey ◽  
Sharmilee Sarkar ◽  
Valentina Teronpi ◽  
Venkata Rajesh Yella ◽  
Aditya Kumar
Keyword(s):  
In Silico ◽  
Pathogenic Bacteria ◽  
Regulatory Regions ◽  
G Quadruplex ◽  
In Cis
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