scholarly journals Structural and functional analysis of the solute-binding protein UspC from Mycobacterium tuberculosis that is specific for amino sugars

Open Biology ◽  
2016 ◽  
Vol 6 (6) ◽  
pp. 160105 ◽  
Author(s):  
Elizabeth Fullam ◽  
Ivan Prokes ◽  
Klaus Fütterer ◽  
Gurdyal S. Besra
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Abc Transporters ◽  
Binding Protein ◽  
Functional Characterization ◽  
Amino Sugar ◽  
Amino Sugars ◽  
Environmental Niche ◽  
Intracellular Infection ◽  
Shift Analysis ◽  
Binding Cleft

Mycobacterium tuberculosis ( Mtb ), the aetiological agent of tuberculosis, has evolved to scavenge nutrients from the confined environment of host macrophages with mycobacterial ATP-binding cassette (ABC) transporters playing a key role in nutrient acquisition. Mtb -UspC (Rv2318) is the solute-binding protein of the essential transporter UspABC, one of four Mtb ABC transporters implicated by homology in sugar acquisition. Herein, we report the structural and functional characterization of Mtb -UspC. The 1.5 Å resolution structure of UspC reveals a two subdomain architecture that forms a highly acidic carbohydrate-substrate binding cleft. This has allowed a distinct preference of Mtb -UspC for amino sugars as determined by thermal shift analysis and solution saturation transfer difference-NMR. Taken together our data support the functional assignment of UspABC as an amino-sugar transporter. Given the limited availability of carbohydrates within the phagosomal environmental niche during Mtb intracellular infection, our studies suggest that UspABC enables Mtb to optimize the use of scarce nutrients during intracellular infection, linking essentiality of this protein to a potential role in recycling components of cell-wall peptidoglycan.

scholarly journals Overexpression and functional characterization of an ABC (ATP-binding cassette) transporter encoded by the genes drrA and drrB of Mycobacterium tuberculosis

2002 ◽  
Vol 367 (1) ◽  
pp. 279-285 ◽  
Author(s):  
Baisakhee Saha CHOUDHURI ◽  
Sanjib BHAKTA ◽  
Rajib BARIK ◽  
Joyoti BASU ◽  
Manikuntala KUNDU ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Abc Transporters ◽  
Efflux Pump ◽  
Functional Characterization ◽  
Atp Binding ◽  
E Coli ◽  
Neutral Compound ◽  
Genes Encoding ◽  
Simultaneous Expression ◽  
Atp Binding Cassette

The genes encoding ATP-binding cassette (ABC) transporters occupy 2.5% of the genome of Mycobacterium tuberculosis. However, none of these putative ABC transporters has been characterized so far. We describe the development of expression systems for simultaneous expression of the ATP-binding protein DrrA and the membrane integral protein DrrB which together behave as a functional doxorubicin efflux pump. Doxorubicin uptake in Escherichia coli or Mycobacterium smegmatis expressing DrrAB was inhibited by reserpine, an inhibitor of ABC transporters. The localization of DrrA to the membrane depended on the simultaneous expression of DrrB. ATP binding was positively regulated by doxorubicin and daunorubicin. At the same time, DrrB appeared to be sensitive to proteolysis when expressed alone in the absence of DrrA. Simultaneous expression of the two polypeptides was essential to obtain a functional doxorubicin efflux pump. Expression of DrrAB in E. coli conferred 8-fold increased resistance to ethidium bromide, a cationic compound. 2′,7′-bis-(2-Carboxyethyl)-5(6)-carboxyfluorescein (BCECF), a neutral compound, also behaved as a substrate of the reconstituted efflux pump. When expressed in M. smegmatis, DrrAB conferred resistance to a number of clinically relevant, structurally unrelated antibiotics. The resistant phenotype could be reversed by verapamil and reserpine, two potent inhibitors of ABC transporters.

scholarly journals Divergent Accumulation of Microbial Residues and Amino Sugars in Loess Soil after Six Years of Different Inorganic Nitrogen Enrichment Scenarios

2021 ◽  
Vol 11 (13) ◽  
pp. 5788
Author(s):  
Dominic Kwadwo Anning ◽  
Zhilong Li ◽  
Huizhen Qiu ◽  
Delei Deng ◽  
Chunhong Zhang ◽  
...  
Keyword(s):  
Plant Biomass ◽  
Inorganic Nitrogen ◽  
Amino Sugar ◽  
N Fertilization ◽  
Biochemical Properties ◽  
Sugar Accumulation ◽  
Amino Sugars ◽  
N Enrichment ◽  
Over Time ◽  
Microbial Residues

Amino sugars are key microbial biomarkers for determining the contribution of microbial residues in soil organic matter (SOM). However, it remains largely unclear as to what extent inorganic nitrogen (N) fertilization can lead to the significant degradation of SOM in alkaline agricultural soils. A six-year field experiment was conducted from 2013 to 2018 to evaluate the effects of chronic N enrichment on microbial residues, amino sugars, and soil biochemical properties under four nitrogen (urea, 46% N) fertilization scenarios: 0 (no-N, control), 75 (low-N), 225 (medium-N), and 375 (high-N) kg N ha−1. The results showed that chronic N enrichment stimulated microbial residues and amino sugar accumulation over time. The medium-N treatment increased the concentration of muramic acid (15.77%), glucosamine (13.55%), galactosamine (18.84%), bacterial residues (16.88%), fungal residues (11.31%), and total microbial residues (12.57%) compared to the control in 2018; however, these concentrations were comparable to the high-N treatment concentrations. The ratio of glucosamine to galactosamine and of glucosamine to muramic acid decreased over time due to a larger increase in bacterial residues as compared to fungal residues. Microbial biomass, soil organic carbon, and aboveground plant biomass positively correlated with microbial residues and amino sugar components. Chronic N enrichment improved the soil biochemical properties and aboveground plant biomass, which stimulated microbial residues and amino sugar accumulation over time.

Molecular and Functional Characterization of One Odorant-Binding Protein Gene OBP3 in Bemisia tabaci (Hemiptera: Aleyrodidae)

2019 ◽  
Author(s):  
Ran Wang ◽  
Yuan Hu ◽  
Peiling Wei ◽  
Cheng Qu ◽  
Chen Luo
Keyword(s):  
Host Plant ◽  
Bemisia Tabaci ◽  
Binding Protein ◽  
Insect Pest ◽  
Critical Role ◽  
Functional Characterization ◽  
Odorant Binding Protein ◽  
Binding Characteristics ◽  
And Function ◽  
Odorant Binding

Abstract Odorant binding proteins (OBPs) of insects play a critical role in chemical perceptions and choice of insect host plant. Bemisia tabaci is a notorious insect pest which can damage more than 600 plant species. In order to explore functions of OBPs in B. tabaci, here we investigated binding characteristics and function of odorant-binding protein 3 in B. tabaci (BtabOBP3). The results indicated that BtabOBP3 shows highly similar sequence with OBPs of other insects, including the typical signature motif of six cysteines. The recombinant BtabOBP3 protein was obtained, and the evaluation of binding affinities to tested volatiles of host plant was conducted, then the results indicated that β-ionone had significantly higher binding to BtabOBP3 among other tested plant volatiles. Furthermore, silencing of BtabOBP3 significantly altered choice behavior of B. tabaci to β-ionone. In conclusion, it has been demonstrated that BtabOBP3 exerts function as one carrier of β-ionone and the results could be contributed to reveal the mechanisms of choosing host plant in B. tabaci.

scholarly journals N-Acetylglucosamine-1-Phosphate Transferase, WecA, as a Validated Drug Target in Mycobacterium tuberculosis

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Stanislav Huszár ◽  
Vinayak Singh ◽  
Alica Polčicová ◽  
Peter Baráth ◽  
María Belén Barrio ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Drug Target ◽  
Functional Characterization ◽  
Drug Candidate ◽  
Content Type ◽  
Chemical Libraries ◽  
Whole Cells ◽  
Encoding Gene

ABSTRACT The mycobacterial phosphoglycosyltransferase WecA, which initiates arabinogalactan biosynthesis in Mycobacterium tuberculosis, has been proposed as a target of the caprazamycin derivative CPZEN-45, a preclinical drug candidate for the treatment of tuberculosis. In this report, we describe the functional characterization of mycobacterial WecA and confirm the essentiality of its encoding gene in M. tuberculosis by demonstrating that the transcriptional silencing of wecA is bactericidal in vitro and in macrophages. Silencing wecA also conferred hypersensitivity of M. tuberculosis to the drug tunicamycin, confirming its target selectivity for WecA in whole cells. Simple radiometric assays performed with mycobacterial membranes and commercially available substrates allowed chemical validation of other putative WecA inhibitors and resolved their selectivity toward WecA versus another attractive cell wall target, translocase I, which catalyzes the first membrane step in the biosynthesis of peptidoglycan. These assays and the mutant strain described herein will be useful for identifying potential antitubercular leads by screening chemical libraries for novel WecA inhibitors.

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