427 research outputs found
Metal ion selectivity and homeostasis: nickel sensing in the human pathogen Helicobacter pylori
No full textHomeostasis of transition metals is crucial for cellular life and is related to the use of metals as cofactors of many enzymes that catalyze an enormous variety of biological reactions. Living organisms elaborated complex and tightly regulated mechanisms to correctly acquire, utilize and allocate metals, neutralizing their harmful potential. In these processes, a fundamental role is played by metal sensor proteins, which are metal-dependent transcription factors that can select, among several intracellular metal ions, the correct metal cofactor, and mediate the specific cellular response by regulating protein expression. Due to the critical function that metal ion homeostasis generally plays in host-pathogen interactions, metal-sensors are considered fundamental virulence factors, because their deletion diminishes or abrogates the survival and colonization of many pathogens. A paradigmatic example is represented by the human pathogen Helicobacter pylori, which relies on the activity of the nickel-dependent enzymes urease and hydrogenase to survive in the acidic stomach mucosa, where it is responsible of several diseases, such as gastric ulcer and cancer. In this bacterium, nickel homeostasis is governed by NikR, a widespread homo-tetrameric transcription factor controlling the expression of proteins involved in nickel trafficking and/or nickel enzymes, in response to different concentrations of intracellular nickel.
In the present study, the modulation of the interaction between Helicobacter pylori NikR (HpNikR) and DNA, exerted by different concentrations of Ni2+ and/or other metal ions, is investigated. In particular, the thermodynamic parameters of the interaction between Ni2+, or other divalent metal ions, and HpNikR are determined by using isothermal titration calorimetry (ITC). The metal ion-dependent capability of HpNikR to bind PureA, the promoter of the urease operon, was investigated by using mobility shift assays, DNAse footprinting and ITC. The results provide a congruent description of the parameters that determine HpNikR binding to metal ions and DNA and yield clear proofs for activation of HpNikR selectively driven by Ni2+, through a protein conformational rearrangement specifically induced by Ni2+ binding (1-2). A general scheme (see Figure) for the nickel selective HpNikR–DNA interaction is proposed.
(1) B. Zambelli, M. Bellucci, A. Danielli, V. Scarlato, S. Ciurli
The Ni2+ binding properties of Helicobacter pylori NikR
Chem. Commun., 2007, 35, pp. 3649 - 3651
(2) B. Zambelli, A. Danielli, S. Romagnoli, P. Neyroz, S. Ciurli, V. Scarlato
High-affinity Ni2+ binding selectively promotes binding of Helicobacter pylori NikR to its target urease promoter
J. Mol. Biol., 2008, 383, pp. 1129 - 114
Selectivity of Metal Ion Binding Specifically Drives the Interaction of a Ni2+-sensor with its operator DNA
No full textTransition metal ions are essential micronutrient for living organisms, being fundamental cofactors of many enzymes that catalyze an enormous variety of biological reactions. At the same time, several metal ions are toxic in their free form for cellular life, demanding a tight regulation of their intracellular trafficking and homeostasis. A paradigmatic example is represented by the human pathogen Helicobacter pylori, which relies on the activity of the nickel-dependent enzymes urease and hydrogenase to survive in the acidic stomach mucosa, where it is responsible of several diseases, such as gastric ulcer and cancer. In this bacterium, homeostasis of essential Ni2+ ions should be tightly controlled to allow growth and colonization. In this processes, NikR, a widespread homo-tetrameric transcription factor controlling the expression of proteins involved in nickel trafficking and/or nickel enzymes, plays a crucial role. The present study investigates the modulation of the interaction between Helicobacter pylori NikR (HpNikR) and DNA, exerted by different concentrations of Ni2+ and/or other metal ions. In particular, the thermodynamic parameters of the interaction between Ni2+, or other divalent metal ions, and HpNikR are determined by using isothermal titration calorimetry (ITC). The metal-dependent capability of HpNikR to bind PureA, the promoter of the urease operon, was determined by using mobility shift assays, DNAse footprinting and ITC. The results provide a congruent description of the factors that determine HpNikR binding to metal ions and DNA and yield clear proofs for activation of HpNikR selectively driven by Ni2+, through a protein conformational rearrangement specifically induced by Ni2+ binding (1-2).
(1) B. Zambelli, M. Bellucci, A. Danielli, V. Scarlato, S. Ciurli; The Ni2+ binding properties of Helicobacter pylori NikR. Chem. Commun., 2007, 35, pp. 3649 - 3651
(2) B. Zambelli, A. Danielli, S. Romagnoli, P. Neyroz, S. Ciurli, V. Scarlato; High-affinity Ni2+ binding selectively promotes binding of Helicobacter pylori NikR to its target urease promoter. J. Mol. Biol., 2008, 383, pp. 1129 - 114
Interaction of Selenoprotein W with 14-3-3 Proteins: A Computational Approach
No full textSelW, a protein containing a selenocysteine (Sec) in a conser- ved Cys-X-X-Sec motif, has been suggested to have an antioxidant role in cell metabolism. SelW is known to specifically interact with different isoforms of 14-3-3 proteins. The latter are involved in several cellular processes such as regulation of the cell cycle, metabolism control, apoptosis, protein trafficking, and gene transcription. 14-3-3 proteins feature a conserved solvent-exposed cysteine residue, in a surface environment prone to induce chemical modifications of the thiol functionality following oxidative stress. The structures of 12 homologous complexes between SelW and 14-3-3 were calculated using sequential alignments, molecular modeling, and docking algorithms guided by known experimental NMR data. These structures reveal the viability of a protein complex in which the conserved Sec residue on SelW approaches the conserved exposed Cys on 14-3-3, making a plausible Sec-Se-S-Cys bond. On the basis of the structural information derived from these calculations, we propose a working hypothesis that entails a role for SelW as a physiological partner of 14-3-3 proteins, able to facilitate a redox-based regulation mechanism
FePO4 NANOPARTILCES AS SOURCE OF NUTRIENTS: EFFECTS ON THE PLANT-SOIL SYSTEM AND EVI-DENCE FOR A SAFE AND SUSTAINABLE NANO-FERTILIZATION
No full textIn the last decade, nanotechnology became a consistent part of the technological progress in modern agriculture, with applications in agri-food technology, nano-biosensoring, plant defence and plant nutrition. Nanomaterials which can provide one or more macro/micro-nutrient to the plant are commonly referred as nanofertilizers. Nevertheless, in the scientific literature there are still few evidence of a successful utilization of nanomaterials as fertilizers. In a previous work, it has been shown that iron phosphate (FePO4) nanoparticles (NPs) can provide either iron (Fe) or phosphate (P) to plants grown in hydroponic. The present study is aimed to highlight the effect of FePO4 NPs used as nanofertilizer in the whole plant-soil system, and to determine if they can represent a safe and effective alternative to conventional fertilizers. To investigate the plant early transcriptomic responses to FePO4 NPs exposure, microarray expression analyses have been performed in maize and cucumber roots grown in hydroponic for 24 hours. Responses of the plants treated with FePO4 NPs were shown to be associated mainly to biotic and abiotic stress, cell wall modulation and regulation of transcription, and triggered a different pattern of responses that was dependent on the nano-size. To evaluate the possibility to apply FePO4 NPs to the soil as fertilizer, two different bare soils were treated. Soil enzyme activities, CO2 respiration and DGGE analyses showed that there was not negative impact of FePO4 NPs onto soil microbial community and metabolic functions, neither toxic effects. Further, FePO4 NPs provided available P in bare soil in respect to triple superphosphate (TSP), even though the efficacy was dependent on the soil characteristics. Moreover, FePO4 NPs represented a source of available P for plant, which grown in soil in controlled condition without significant differences in respect to TSP, although P availability in the bare soil resulted lower for NPs than TSP. Microbial community associated to rhizosphere was not negatively affected by NPs and a stimulatory effect on enzyme activity was observed. In this work it was shown that FePO4 NPs can be applied to the soil without any negative consequence for the environment, enhancing plant growth and providing nutrients. These results encourage the hypothesis that the nanoparticulate nature of fertilizers could contribute to rationalize the chemical inputs in agriculture and increasing nutrient use efficienc
POSSIBILE ASSORBIMENTO RADICALE DI AGENTI VIRALI DA ACQUE IRRIGUE CONTAMINATE
No full textPS13.51 - 383
POSSIBILE ASSORBIMENTO RADICALE DI AGENTI
VIRALI DA ACQUE IRRIGUE CONTAMINATE
Verani M.*[1], Ciurli A.[2], Carducci A.[1]
[1]Laboratorio di Igiene e Virologia Ambientale – Dipartimento
di Biologia – Università di Pisa ~ Pisa
[2]Dipartimento di Biologia delle Piante Agrarie – Università
di Pisa ~ Pisa
OBIETTIVI: I vegetali a foglia larga come l’insalata,
sono particolarmente a rischio quando la loro irrigazione
viene effettuata con acqua contaminata
come i liquami parzialmente depurati nei quali e di
solito abbondante la presenza di agenti biologici di
origine fecale, tra cui i virus. E’ ben nota la possibilita
di una contaminazione superficiale dei vegetali
attraverso l'irrigazione a spruzzo, ma recentemente
e stato ipotizzato, e talora evidenziato sperimentalmente,
l’assorbimento di tali agenti attraverso le
radici e quindi il loro passaggio alle foglie quando
acque contaminate vengano utilizzate per la subirrigazione
o per colture idroponiche. In tal caso il lavaggio
superficiale sarebbe insufficiente a rimuovere
la contaminazione. Allo scopo di dimostrare tale
adsorbimento e di stimare il livello di contaminazione
delle acque da considerare a rischio e stato
intrapreso uno studio sperimentale in colture idroponiche
di insalata (Lactuca sativa) addizionate di
virus Coxsackies B2.
METODI: Le piantine d’insalata sono state coltivate
in apposite vasche contenenti soluzione nutritiva
contaminata con Coxsakievirus B2 a 3 diluizioni note.
Per valutare l’eventuale penetrazione del virus
attraverso le radici sono state analizzate, per 4
giorni consecutivi, sia le foglie sia l’acqua delle vasche,
mediante una prima fase di eluizioneconcentrazione
con tecniche gia precedentemente
testate e successiva rilevazione virale con metodologie
colturali e molecolari.
RISULTATI: Le prove hanno evidenziato la presenza
di virus (anche su colture cellulari e quindi infettivo)
nelle foglie gia dal primo giorno dopo la contaminazione
con una concentrazione crescente per tutta la
durata dell’esperimento. Contemporaneamente e
stata riscontrato un decadimento piuttosto rapido
del titolo virale nelle acque con entrambi i metodi
di analisi.
CONCLUSIONI: Tali risultati indicano che quando le
radici sono a contatto con acque contaminate a
concentrazione virale superiore a 10^3 GC/ml e
possibile avere penetrazione a livello delle foglie.
Poiche tale valore puo ritrovarsi frequentemente
nei reflui anche parzialmente depurati, e importante
che la qualita virologica delle acque di riuso agricolo
venga valutata prima dell’utilizzo
Intrinsic Fluorescence of Intrinsically Disordered Proteins
No full textResolution of the intrinsic emission properties of a protein by different fluorescence spectroscopy techniques is an invaluable tool to detect and characterize its structural architecture and conformational changes under different experimental conditions. Indeed, the multidimensional character of fluorescence can provide information on local chemical features, on solvent diffusional processes, and on rotational movements of peptide chains or whole proteins. Here, we describe the details of quenching fluorescence experiments and how to correlate the results to the peculiar structural information on the organization of intrinsically disordered proteins (IDPs)
Redox activity of caffeic acid towards iron (III) complexed in a polygalacturonate network
No full textThe transfer of several metal ions from the soil to the plant absorbing cells is mediated principally by organic molecules of low molecular weight with complexing and reducing activity, among which caffeic acid (CAF) is particularly important. Here we report the results of a survey which deals with the oxidation of CAF by the Fe(III) ions bound to a polygalacturonate network (Fe(III)-PGA network). The interaction between Fe(III) and CAF was studied by using Fe(III)-PGA networks equilibrated in the 2.4-7.0 pH range by means of kinetic and spectroscopic methods. The reducing power was found to depend on the nature of the Fe(III)-PGA network complexes: when the ferric ion was complexed only by the PGA carboxylic groups, a high redox activity was observed, whereas the Fe(III) reduction was found to be lower when a hydroxylic group was inserted in the Fe(III) coordination sphere. The iron complexed in the network was protected from hydrolysis reactions, as shown by the high pH values at which its reduction occurred. Two different fractions of Fe(II) produced were identified, one diffusible and another exchangeable with CaCl2 6.0 mM. The existence of the exchangeable form was attributed to the electrostatic interaction of the Fe(II) ions with the carboxylate groups of the fibrils and with the degradation products of CAE The arrangement of the fibrils was altered following the substitution of Ca(II) by Fe(III) ions and was restored following the seduction of Fe(III) by CAF
Commercial advantages on basil architecture by ultraviolet-B irradiation
No full textSweet basil (Ocimum basilicum L.) is one of the most important herbs widely used for its medicinal properties and as food ingredient. The marketing of this product highlights the problem that these plants have long and slender stems, which are easy to break off and thus making difficult their market distribution. In this work, two cultivars of basil (Genovese and Profumo) at the adequate development stage for sale were used. We evaluated the effect of supplemental ultraviolet (UV)-B irradiation (15 W m-2; 3 h day-1) on plant growth and market quality. Both cultivars of basil plants under UV-B irradiation resulted in increased leaf size and biomass, and decreased shoot length in comparison to that of under control growth conditions. These results indicate that the application of UV-B irradiation beneficially influenced plant architecture in basil improving their greenhouse production for fresh market
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