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Ítem Acceso Abierto Alkylhalovinylboranes: a new class of Diels-Alder dienophiles(Royal Society of Chemistry, 2018-10-02) Pisano, Pablo Luis; Pellegrinet, Silvina Carla; https://orcid.org/0000-0002-6368-9234; https://orcid.org/0000-0001-6854-7377The Diels–Alder reactions of alkylhalovinylboranes have been investigated theoretically and experimentally. Alkylhalovinylboranes presented higher reactivity than the corresponding dialkylvinylboranes. Although endo/exo selectivities were high for the reactions with cyclopentadiene, facial selectivities for the chiral analogues were low. Our results demonstrate that the replacement of an alkyl group on the boron atom by a halogen increases the dienophilicity considerably.Ítem Acceso Abierto Pleiotropic effect of AccD5 and AccE5 depletion in acyl-coenzyme a carboxylase activity and in lipid biosynthesis in mycobacteria(Public Library of Science, 2014-06-20) Bazet Lyonnet, Bernardo; Diacovich, Lautaro; Cabruja, Matías Ezequiel; Bardou, Fabienne; Quémard, Annaïk; Gago, Gabriela; Gramajo, Hugo CesarMycobacteria contain a large variety of fatty acids which are used for the biosynthesis of several complex cell wall lipids that have been implicated in the ability of the organism to resist host defenses. The building blocks for the biosynthesis of all these lipids are provided by a fairly complex set of acyl-CoA carboxylases (ACCases) whose subunit composition and roles within these organisms have not yet been clearly established. Previous biochemical and structural studies provided strong evidences that ACCase 5 from Mycobacterium tuberculosis is formed by the AccA3, AccD5 and AccE5 subunits and that this enzyme complex carboxylates acetyl-CoA and propionyl-CoA with a clear substrate preference for the latest. In this work we used a genetic approach to unambiguously demonstrate that the products of both accD5 and accE5 genes are essential for the viability of Mycobacterium smegmatis. By obtaining a conditional mutant on the accD5-accE5 operon, we also demonstrated that the main physiological role of this enzyme complex was to provide the substrates for fatty acid and mycolic acid biosynthesis. Furthermore, enzymatic and biochemical analysis of the conditional mutant provided strong evidences supporting the notion that AccD5 and/or AccE5 have an additional role in the carboxylation of long chain acylCoA prior to mycolic acid condensation. These studies represent a significant step towards a better understanding of the roles of ACCases in mycobacteria and confirm ACCase 5 as an interesting target for the development of new antimycobacterial drugs.Ítem Acceso Abierto Metal-mediated synthesis of pyrrolines(Royal Society of Chemistry, 2019-02-27) Medran, Noelia Soledad; La Venia, Agustina; Testero, Sebastián A.; https://orcid.org/0000-0002-0185-5751The five-membered, nitrogen-containing pyrroline ring is a privileged structure. This ring is present in many bioactive compounds from natural sources. Pyrrolines—the dihydro derivatives of pyrroles—have three structural isomer classes, depending on the location of the double bond: 1-pyrrolines (3,4-dihydro-2H-pyrroles), 2-pyrrolines (2,3-dihydro-1H-pyrroles) and 3-pyrrolines (2,5-dihydro-1H-pyrroles). This review aims to describe the latest advances for the synthesis of pyrrolines by transition metal-catalyzed cyclizations. Only reactions in which the pyrroline ring is formed by metal promotion are described. Transformations of the pyrroline ring in other heterocycles, and the structural manipulations of the pyrroline itself are not discussed. The review is organized into three parts, each covering the metal-mediated synthesis of the three pyrroline isomers. Each part is subdivided according to the metal involved, and concludes with a brief description of notable biological activities within the class.Ítem Acceso Abierto Structural requirements for the antifungal activities of natural drimane sesquiterpenes and analogues, supported by conformational and electronic studies(MDPI, 2013-02-05) Derita, Marcos Gabriel; Montenegro, Iván; Garibotto, Francisco; Enriz, Ricardo Daniel; Cuellar Fritis, Mauricio; Zacchino, Susana; https://orcid.org/0000-0002-7112-5500Seventeen drimanes including polygodial (1), isopolygodial (2), drimenol (3) and confertifolin (4) obtained from natural sources and the semi-synthetic derivatives 5–17 obtained from 1–3, were evaluated in vitro for antifungal properties against a unique panel of fungi with standardized procedures by using two end-points, MIC100 and MIC50. A SAR analysis of the whole series, supported by conformational and electronic studies, allowed us to show that the Δ7,8 -double bond would be one of the key structural features related to the antifungal activity. The MEPs obtained for active compounds exhibit a clear negative minimum value (deep red zone) in the vicinity of the Δ7,8 -double bond, which is not present in the inactive ones. Apart of this negative zone, a positive region (deep blue) appears in 1, which is not observed either in its epimer 2 nor in the rest of the active compounds. The LogP of active compounds varies between 2.33 and 3.84, but differences in MICs are not correlated with concomitant variations in LogP values.Ítem Acceso Abierto A penicillin derivative exerts an anti-metastatic activity in melanoma cells through the downregulation of integrin αvβ3 and Wnt/β-Catenin pathway(Frontiers Media, 2020-02-25) Barrionuevo, Elizabeth; Cayrol, María Florencia; Cremaschi, Graciela Alicia; Cornier, Patricia Griselda; Boggián, Dora Bernarda; Delpiccolo, Carina M. L.; Mata, Ernesto Gabino; Roguin, Leonor P.; Blank, Viviana C.The synthetic triazolylpeptidyl penicillin derivative, named TAP7f, has been previously characterized as an effective antitumor agent in vitro and in vivo against B16-F0 melanoma cells. In this study, we investigated the anti-metastatic potential of this compound on highly metastatic murine B16-F10 and human A375 melanoma cells. We found that TAP7f inhibited cell adhesion, migration and invasion in a dose-dependent manner. Additionally, we demonstrated that TAP7f downregulated integrin αvβ3 expression and Wnt/β-catenin pathway, a signaling cascade commonly related to tumor invasion and metastasis. Thus, TAP7f reduced both the enzymatic activity and the expression levels of matrix-metalloproteinases-2 and -9 in a time dependent manner. Moreover, TAP7f inhibited the expression of the transcription factor Snail and the mesenchymal markers vimentin, and N-cadherin, and up-regulated the expression of the epithelial marker E-cadherin, suggesting that the penicillin derivative affects epithelial–mesenchymal transition. Results obtained in vitro were supported by those obtained in a B16-F10-bearing mice metastatic model, that showed a significant TAP7f inhibition of lung metastasis. These findings suggest the potential of TAP7f as a chemotherapeutic agent for the treatment of metastatic melanoma.Ítem Acceso Abierto High cell density production of multimethyl-branched long-chain esters in Escherichia coli and determination of their physicochemical properties(BMC (part of Springer Nature), 2016-10-14) Menendez Bravo, Simón M.; Roulet, Julia; Sabatini, Martín; Comba, Santiago; Dunn, Robert; Gramajo, Hugo Cesar; Arabolaza, Ana LorenaBackground: microbial synthesis of oleochemicals derived from native fatty acid (FA) metabolism has presented significant advances in recent years. Even so, native FA biosynthetic pathways often provide a narrow variety of usually linear hydrocarbons, thus yielding end products with limited structural diversity. To overcome this limitation, we took advantage of a polyketide synthase-based system from Mycobacterium tuberculosis and developed an Escherichia coli platform with the capacity to synthesize multimethyl-branched long-chain esters (MBE) with novel chemical structures. Results: with the aim to initiate the characterization of these novel waxy compounds, here, we describe the chassis optimization of the MBE producer E. coli strain for an up-scaled oil production. By carrying out systematic metabolic engineering, we improved the final titer to 138.1 ± 5.3 mg MBE L−1 in batch cultures. Fed-batch microbial fermentation process was also optimized achieving a maximum yield of 790.2 ± 6.9 mg MBE L−1 with a volumetric productivity of 15.8 ± 1.1 mg MBE (L h)−1. Purified MBE oil was subjected to various physicochemical analyses, including differential scanning calorimetry (DSC) and pressurized-differential scanning calorimetry (P-DSC) studies. Conclusions: the analysis of the pour point, DSC, and P-DSC data obtained showed that bacterial MBE possess improved cold flow properties than several plant oils and some chemically modified derivatives, while exhibiting high oxidation stability at elevated temperatures. These encouraging data indicate that the presence of multiple methyl branches in these novel esters, indeed, conferred favorable properties which are superior to those of linear esters.Ítem Acceso Abierto Engineering a Streptomyces coelicolor biosynthesis pathway into Escherichia coli for high yield triglyceride production(BMC, 2014-12) Comba, Santiago; Sabatini, Martín; Menendez Bravo, Simón M.; Arabolaza, Ana Lorena; Gramajo, Hugo CesarBackground: Microbial lipid production represents a potential alternative feedstock for the biofuel and oleochemical industries. Since Escherichia coli exhibits many genetic, technical, and biotechnological advantages over native oleaginous bacteria, we aimed to construct a metabolically engineered E. coli strain capable of accumulating high levels of triacylglycerol (TAG) and evaluate its neutral lipid productivity during high cell density fed-batch fermentations. Results: The Streptomyces coelicolor TAG biosynthesis pathway, defined by the acyl-CoA:diacylglycerol acyltransferase (DGAT) Sco0958 and the phosphatidic acid phosphatase (PAP) Lppβ, was successfully reconstructed in an E. coli diacylglycerol kinase (dgkA) mutant strain. TAG production in this genetic background was optimized by increasing the levels of the TAG precursors, diacylglycerol and long-chain acyl-CoAs. For this we carried out a series of stepwise optimizations of the chassis by 1) fine-tuning the expression of the heterologous SCO0958 and lppβ genes, 2) overexpression of the S. coelicolor acetyl-CoA carboxylase complex, and 3) mutation of fadE, the gene encoding for the acyl-CoA dehydrogenase that catalyzes the first step of the β-oxidation cycle in E. coli. The best producing strain, MPS13/pET28-0958 ACC/pBAD-LPPβ rendered a cellular content of 4.85% cell dry weight (CDW) TAG in batch cultivation. Process optimization of fed-batch fermentation in a 1-L stirred-tank bioreactor resulted in cultures with an OD600nm of 80 and a product titer of 722.1 mg TAG L-1 at the end of the process. Conclusions: This study represents the highest reported fed-batch productivity of TAG reached by a model non-oleaginous bacterium. The organism used as a platform was an E. coli BL21 derivative strain containing a deletion in the dgkA gene and containing the TAG biosynthesis genes from S. coelicolor. The genetic studies carried out with this strain indicate that diacylglycerol (DAG) availability appears to be one of the main limiting factors to achieve higher yields of the storage compound. Therefore, in order to develop a competitive process for neutral lipid production in E. coli, it is still necessary to better understand the native regulation of the carbon flow metabolism of this organism, and in particular, to improve the levels of DAG biosynthesis.Ítem Acceso Abierto HrpE, the major component of the Xanthomonas type three protein secretion pilus, elicits plant immunity responses(Springer Nature, 2018-06-29) Gottig, Natalia; Vranych, Cecilia Verónica; Sgro, Germán Gustavo; Piazza, Ainelén; Ottado, JorgelinaLike several pathogenic bacteria, Xanthomonas infect host plants through the secretion of effector proteins by the Hrp pilus of the Type Three Protein Secretion System (T3SS). HrpE protein was identified as the major structural component of this pilus. Here, using the Xanthomonas citri subsp. citri (Xcc) HrpE as a model, a novel role for this protein as an elicitor of plant defense responses was found. HrpE triggers defense responses in host and non-host plants revealed by the development of plant lesions, callose deposition, hydrogen peroxide production and increase in the expression levels of genes related to plant defense responses. Moreover, pre-infiltration of citrus or tomato leaves with HrpE impairs later Xanthomonas infections. Particularly, HrpE C-terminal region, conserved among Xanthomonas species, was sufficient to elicit these responses. HrpE was able to interact with plant Glycine-Rich Proteins from citrus (CsGRP) and Arabidopsis (AtGRP-3). Moreover, an Arabidopsis atgrp-3 knockout mutant lost the capacity to respond to HrpE. This work demonstrate that plants can recognize the conserved C-terminal region of the T3SS pilus HrpE protein as a danger signal to defend themselves against Xanthomonas, triggering defense responses that may be mediated by GRPs.Ítem Acceso Abierto May the force (field) be with you: on the importance of conformational searches in the prediction of NMR chemical shifts(MDPI, 2022-11-08) Cuadrado, Cristina; Hernández Daranas, Antonio; Sarotti, Ariel Marcelo; https://orcid.org/0000-0001-9282-1575NMR data prediction is increasingly important in structure elucidation. The impact of force field selection was assessed, along with geometry and energy cutoffs. Based on the conclusions, we propose a new approach named mix-J-DP4, which provides a remarkable increase in the confidence level of complex stereochemical assignments—100% in our molecular test set—with a very modest increment in computational cost.Ítem Acceso Abierto Botanical control of citrus green mold and peach brown rot on fruits assays using a persicaria acuminata phytochemically characterized extract(MDPI, 2021-02-24) Di Liberto, Melina G.; Seimandi, Gisela; Fernández, Laura; Ruiz, Verónica; Svetaz, Laura Andrea; Derita, Marcos Gabriel; https://orcid.org/0000-0002-0162-2771; https://orcid.org/0000-0001-8148-936XPersicaria acuminata (Polygonaceae) is a perennial herb that grows in the central area of Argentina and it is commonly used by native populations to heal infected wounds and other conditions related to fungal infections. In this article, we explored the in vitro antifungal activity of its ethyl acetate extract against a panel of three fruit phytopathogenic fungi including: Penicillium digitatum, P. italicum, and Monilinia fructicola. The sesquiterpenes isolated from the extract were also evaluated against these strains, demonstrating that the dialdehyde polygodia was the responsible for this activity. In order to encourage the use of the extract rather than the pure compound, we displayed ex vivo assays using fresh oranges and peaches inoculated with P. digitatum and M. fructicola, respectively, and subsequently treated by immersion with an extract solution of 250 and 62.5 µg/mL, respectively. There were no statistically significant differences between the treatments with commercial fungicides and the extract over the control of both fruit rots. The concentration of the active compound present in the extract used on fruit experiments was determined by Gas Chromatography-Mass Spectroscopy. Finally, cytotoxicity evaluation against Huh7 cells showed that P. acuminata extract was less cytotoxic than the commercial fungicides at the assayed concentrations. After these findings we could conclude that a chemically characterized extract of P. acuminata should be further developed to treat fungal diseases in fruits from an agro-ecological model.Ítem Acceso Abierto Identification and physiological characterization of phosphatidic acid phosphatase enzymes involved in triacylglycerol biosynthesis in Streptomyces coelicolor(BMC, 2013-01-29) Comba, Santiago; Menendez Bravo, Simón M.; Arabolaza, Ana Lorena; Gramajo, Hugo CesarBackground: Phosphatidic acid phosphatase (PAP, EC 3.1.3.4) catalyzes the dephosphorylation of phosphatidate yielding diacylglycerol (DAG), the lipid precursor for triacylglycerol (TAG) biosynthesis. Despite the importance of PAP activity in TAG producing bacteria, studies to establish its role in lipid metabolism have been so far restricted only to eukaryotes. Considering the increasing interest of bacterial TAG as a potential source of raw material for biofuel production, we have focused our studies on the identification and physiological characterization of the putative PAP present in the TAG producing bacterium Streptomyces coelicolor. Results: We have identified two S. coelicolor genes, named lppα (SCO1102) and lppβ (SCO1753), encoding for functional PAP proteins. Both enzymes mediate, at least in part, the formation of DAG for neutral lipid biosynthesis. Heterologous expression of lppα and lppβ genes in E. coli resulted in enhanced PAP activity in the membrane fractions of the recombinant strains and concomitantly in higher levels of DAG. In addition, the expression of these genes in yeast complemented the temperature-sensitive growth phenotype of the PAP deficient strain GHY58 (dpp1lpp1pah1). In S. coelicolor, disruption of either lppα or lppβ had no effect on TAG accumulation; however, the simultaneous mutation of both genes provoked a drastic reduction in de novo TAG biosynthesis as well as in total TAG content. Consistently, overexpression of Lppα and Lppβ in the wild type strain of S. coelicolor led to a significant increase in TAG production. Conclusions: The present study describes the identification of PAP enzymes in bacteria and provides further insights on the genetic basis for prokaryotic oiliness. Furthermore, this finding completes the whole set of enzymes required for de novo TAG biosynthesis pathway in S. coelicolor. Remarkably, the overexpression of these PAPs in Streptomyces bacteria contributes to a higher productivity of this single cell oil. Altogether, these results provide new elements and tools for future cell engineering for next-generation biofuels production.Ítem Acceso Abierto Efficient Buchwald–Hartwig and nitrene-mediated five-membered ring closure approaches to the total synthesis of quindoline. Unexpected direct conversion of a nitro group into a phosphazene(Royal Society of Chemistry, 2023-05-05) Thobokholt, Elida N.; Simonetti, Sebastián Osvaldo; Kaufman, Teodoro Saúl; Larghi, Enrique Leandro; Bracca, Andrea Beatriz Juana; https://orcid.org/0000-0001-5780-6433; https://orcid.org/0000-0003-4236-7380; https://orcid.org/0000-0003-3173-2178; https://orcid.org/0000-0002-9791-4831; https://orcid.org/0000-0003-2647-475XTwo total syntheses of quindoline, which take place through the intermediacy of 3-nitroquinoline derivatives, are reported. The general synthetic sequence involves construction of the latter by mechanochemical condensation of benzaldehydes with 2-amino-nitrostyrene, followed either by reduction of the nitro group of the heterocycle and Buchwald–Hartwig cyclization or by a nitrenemediated cyclization under solventless conditions. Use of PPh3 to generate the nitrene resulted in the unprecedented formation of a phosphazene in place of quindoline. This unexpected transformation was explained by means of DFT computations.Ítem Acceso Abierto A coiled coil switch mediates cold sensing by the thermosensory protein DesK(Wiley, 2015-10-08) Saita, Emilio Adolfo; Abriata, Luciano Andrés; Tsai, Yi-Ting; Trajtenberg, Felipe; Lemmin, Thomas; Buschiazzo, Alejandro; Dal Peraro, Matteo; De Mendoza, Diego; Albanesi, DanielaThe thermosensor histidine kinase DesK from Bacillus subtilis senses changes in membrane fluidity initiating an adaptive response. Structural changes in DesK have been implicated in transmembrane signaling, but direct evidence is still lacking. On the basis of structure-guided mutagenesis, we now propose a mechanism of DesK-mediated signal sensing and transduction. The data indicate that stabilization/destabilization of a 2-helix coiled coil, which connects the transmembrane sensory domain of DesK to its cytosolic catalytic region, is crucial to control its signaling state. Computational modeling and simulations reveal couplings between protein, water and membrane mechanics. We propose that membrane thickening is the main driving force for signal sensing and that it acts by inducing helix stretching and rotation prompting an asymmetric kinase-competent state. Overall, the known structural changes of the sensor kinase, as well as further dynamic rearrangements that we now predict, consistently link structure determinants to activity modulation.Ítem Acceso Abierto The pleiotropic transcriptional regulator NlpR contributes to the modulation of nitrogen metabolism, lipogenesis and triacylglycerol accumulation in oleaginous rhodococci(Wiley, 2016-11-25) Hernández, Martín Alejandro; Lara, María Julia; Gago, Gabriela; Gramajo, Hugo Cesar; Álvarez, Héctor ManuelThe regulatory mechanisms involved in lipogenesis and triacylglycerol (TAG) accumulation are largely unknown in oleaginous rhodococci. In this study a regulatory protein (here called NlpR: Nitrogen lipid Regulator), which contributes to the modulation of nitrogen metabolism, lipogenesis and triacylglycerol accumulation in oleaginous rhodococci was identified. Under nitrogen deprivation conditions, in which TAG accumulation is stimulated, the nlpR gene was significantly upregulated, whereas a significant decrease of its expression and TAG accumulation occurred when cerulenin was added. The nlpR disruption negatively affected the nitrate/nitrite reduction as well as lipid biosynthesis under nitrogen-limiting conditions. In contrast, its overexpression increased TAG production during cultivation of cells in nitrogen-rich media. A putative ‘NlpR-binding motif’ upstream of several genes related to nitrogen and lipid metabolisms was found. The nlpR disruption in RHA1 strain led to a reduced transcription of genes involved in nitrate/nitrite assimilation, as well as in fatty acid and TAG biosynthesis. Purified NlpR was able to bind to narK, nirD, fasI, plsC and atf3 promoter regions. It was suggested that NlpR acts as a pleiotropic transcriptional regulator by activating of nitrate/nitrite assimilation genes and others genes involved in fatty acid and TAG biosynthesis, in response to nitrogen deprivation.Ítem Acceso Abierto Furo[3,2-c]coumarins carrying carbon substituents at C-2 and/or C-3. Isolation, biological activity, synthesis and reaction mechanisms(Royal Society of Chemistry, 2020) Cortés, Iván; Cala, Javier L.; Bracca, Andrea Beatriz Juana; Kaufman, Teodoro Saúl; http://orcid.org/0000-0002-8069-2793; http://orcid.org/0000-0003-2647-475X; http://orcid.org/0000-0003-3173-2178The isolation, biological activity and synthesis of natural furo[3,2-c]coumarins are presented, covering mainly the developments in the last 35 years. The most relevant approaches toward the synthesis of 2- substituted, 3-substituted and 2,3-disubstituted heterocycles are also discussed, with emphasis on the efficiency of the processes and their mechanisms.Ítem Acceso Abierto Crystal structure of cis-1-phenyl-8-(pyridin-2-yl-methyl)dibenzo[1,2-c:2,1-h]-2,14-dioxa-8-aza-1-borabicyclo[4.4.0]deca-3,8-diene(International Union of Crystallography, 2017-12-01) Ledesma, Gabriela N.; Signorella, Sandra R.; Back, Davi; Schulz Lang, ErnestoThe title compound, C26H23BN2O2, was obtained as by product during synthetic attempts of a complexation reaction between the tripodal ligand H2L [N,N-bis(2-hydroxybenzyl)(pyridin-2-yl)methylamine] and manganese(III) acetate in the presence of NaBPh4. The isolated B-phenyl dioxazaborocine contains an N→B dative bond with a cis conformation. In the crystal, C—H⋯O hydrogen bonds define chains parallel to the b-axis direction. A comparative analysis with other structurally related derivatives is also included, together with a rationalization of the unexpected production of this zwitterionic heterocycle.Ítem Acceso Abierto Antifungal activity and toxicity studies of flavanones isolated from Tessaria dodoneifolia aerial parts(Elsevier, 2020) Soberón, José R.; Sgariglia, Melina A.; Carabajal Torrez, José A.; Aguilar, Franco A.; Pero, Edgardo J.I.; Sampietro, Diego A.; Fernández de Luco, Julia; Labadie, Guillermo RobertoTessaria dodoneifolia [Asteraceae] is traditionally employed in Northwestern Argentina for fungal infections treatment. We report the antifungal activity guided isolation and identification of substances from aerial parts of this species, both individually and in combination with fluconazole (FLU), against Candida albicans strains. Two antifungal flavanones were identified as naringenin (NAR) and pinocembrin (PIN). These compounds could individually inhibit the growth of C. albicans strains. Combinations of NAR and PIN with FLU were synergistic against the FLU resistant and sensitive C. albicans strains. Genotoxic and cytotoxic evaluations were also performed. NAR, PIN and their combinations with FLU did not have a genotoxic effect on Bacillus subtilis rec strains. Finally, these compounds did not show cytotoxicity at concentrations below 80 μg/mL.Ítem Acceso Abierto Mycobacterium tuberculosis FasR senses long fatty acyl-CoA through a tunnel and a hydrophobic transmission spine(Springer Nature, 2020-07-24) Lara, María Julia; Diacovich, Lautaro; Trajtenberg, Felipe; Larrieux, Nicole; Malchiodi, Emilio L.; Fernández, Marisa M.; Gago, Gabriela; Gramajo, Hugo Cesar; Buschiazzo, Alejandro; https://orcid.org/0000-0002-3339-0100; https://orcid.org/0000-0003-0427-5549; https://orcid.org/0000-0001-7501-3330; https://orcid.org/0000-0001-7668-0119; https://orcid.org/0000-0002-2509-6526Mycobacterium tuberculosis is a pathogen with a unique cell envelope including very long fatty acids, implicated in bacterial resistance and host immune modulation. FasR is a TetR-like transcriptional activator that plays a central role in sensing mycobacterial long-chain fatty acids and regulating lipid biosynthesis. Here we disclose crystal structures of M. tuberculosis FasR in complex with acyl effector ligands and with DNA, uncovering its molecular sensory and switching mechanisms. A long tunnel traverses the entire effector-binding domain, enabling long fatty acyl effectors to bind. Only when the tunnel is entirely occupied, the protein dimer adopts a rigid configuration with its DNA-binding domains in an open state, leading to DNA dissociation. The protein-folding hydrophobic core connects the two domains, and is completed into a continuous spine when the effector binds. Such a transmission spine is conserved in a large number of TetR-like regulators, offering insight into effector-triggered allosteric functional control.Ítem Acceso Abierto XacFhaB adhesin, an important Xanthomonas citri ssp. citri virulence factor, is recognized as a pathogen-associated molecular pattern(BSPP and Wiley, 2016-11-15) Garavaglia, Betiana Soledad; Zimaro, Tamara; Abriata, Luciano Andrés; Ottado, Jorgelina; Gottig, NataliaAdhesion to host tissue is one of the key steps of the bacterial pathogenic process. Xanthomonas citri ssp. citri possesses a non-fimbrial adhesin protein, XacFhaB, required for bacterial attachment, which we have previously demonstrated to be an important virulence factor for the development of citrus canker. XacFhaB is a 4753-residue-long protein with a predicted β-helical fold structure, involved in bacterial aggregation, biofilm formation and adhesion to the host. In this work, to further characterize this protein and considering its large size, XacFhaB was dissected into three regions based on bioinformatic and structural analyses for functional studies. First, the capacity of these protein regions to aggregate bacterial cells was analysed. Two of these regions were able to form bacterial aggregates, with the most amino-terminal region being dispensable for this activity. Moreover, XacFhaB shows features resembling pathogen-associated molecular patterns (PAMPs), which are recognized by plants. As PAMPs activate plant basal immune responses, the role of the three XacFhaB regions as elicitors of these responses was investigated. All adhesin regions were able to induce basal immune responses in host and non-host plants, with a stronger activation by the carboxyl-terminal region. Furthermore, pre-infiltration of citrus leaves with XacFhaB regions impaired X. citri ssp. citri growth, confirming the induction of defence responses and restraint of citrus canker. This work reveals that adhesins from plant pathogens trigger plant defence responses, opening up new pathways for the development of protective strategies for disease control.Ítem Acceso Abierto Langmuir-Blodgett monolayers holding a wound healing active compound and its effect in cell culture. A model for the study of surface mediated drug delivery systems(Elsevier, 2021-03-14) Fernández, Luciana Andrea; Reviglio, Ana Lucía; Heredia, Daniel Alejandro; Morales, Gustavo Marcelo; Santo, Marisa Rosana; Otero, Luis Alberto; Alustiza, Fabrisio Eduardo; Liaudat, Ana Cecilia; Bosch, Pablo; Larghi, Enrique Leandro; Bracca, Andrea Beatriz Juana; Kaufman, Teodoro SaúlLangmuir and Langmuir-Blodgett films holding a synthetic bioinspired wound healing active compound were used as drug-delivery platforms. Palmitic acid Langmuir monolayers were able to incorporate 2-methyltriclisine, a synthetic Triclisine derivative that showed wound healing activity. The layers proved to be stable and the nanocomposites were transferred to solid substrates. Normal human lung cells (Medical Research Council cell strain 5, MRC-5) were grown over the monomolecular Langmuir-Blodgett films that acted as a drug reservoir and delivery system. The proliferation and migration of the cells were clearly affected by the presence of 2 methyltriclisine in the amphiphilic layers. The methodology is proposed as a simple and reliable model for the study of the effects of bioactive compounds over cellular cultures.