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Articles de revues sur le sujet "HrpA protein"
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Frederick, Reid D., Musharaf Ahmad, Doris R. Majerczak, Angel S. Arroyo-Rodríguez, Shulamit Manulis et David L. Coplin. « Genetic Organization of the Pantoea stewartii subsp. stewartii hrp Gene Cluster and Sequence Analysis of the hrpA, hrpC, hrpN, and wtsE Operons ». Molecular Plant-Microbe Interactions® 14, no 10 (octobre 2001) : 1213–22. http://dx.doi.org/10.1094/mpmi.2001.14.10.1213.
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The hrp/wts gene cluster of Pantoea stewartii subsp. stewartii is required for pathogenicity on sweet corn and the ability to elicit a hypersensitive response (HR) in tobacco. Site-directed transposon mutagenesis and nucleotide sequencing were used to identify hrp/wts genes within the left 20 kb of this cluster. Seventeen open reading frames (ORFs) comprise seven genetic complementation groups. These ORFs share homology with hrp and dsp genes from Erwinia amylovora, Erwinia chrysanthemi, and Pseudomonas syringae pathovars and have been designated, in map order, wtsF, wtsE, hrpN, hrpV, hrpT, hrcC, hrpG, hrpF, hrpE, hrpD, hrcJ, hrpB, hrpA, hrpS, hrpY, hrpX, and hrpL. Putative hrp consensus promoter sequences were identified upstream of hrpA, hrpF, hrpN, and wtsE. Expression of the hrpA, hrpC, and wtsE operons was regulated by HrpS. Transposon mutations in all of the hrp operons abolished pathogenicity and HR elicitation, except for the hrpN and hrpV mutants, which were still pathogenic. hrpS, hrpXY, and hrpL regulatory mutations abolished HrpN synthesis, whereas secretory mutations in the hrpC, hrpA, and hrpJ operons permitted intracellular HrpN synthesis. wtsEF mutants were not pathogenic but still produced HrpN and elicited the HR. wtsE encodes a 201-kDa protein that is similar to DspE in E. amylovora and AvrE in P. syringae pv. tomato, suggesting that this protein is a major virulence factor involved in the elicitation of water-soaked lesions.
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Schmitt, Corinna, David Turner, Maria Boesl, Marion Abele, Matthias Frosch et Oliver Kurzai. « A Functional Two-Partner Secretion System Contributes to Adhesion of Neisseria meningitidis to Epithelial Cells ». Journal of Bacteriology 189, no 22 (14 septembre 2007) : 7968–76. http://dx.doi.org/10.1128/jb.00851-07.
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ABSTRACT Neisseria meningitidis is a frequent commensal of the human nasopharynx causing severe invasive infections in rare cases. A functional two-partner secretion (TPS) system in N. meningitidis, composed of the secreted effector protein HrpA and its cognate transporter HrpB, is identified and characterized in this study. Although all meningococcal strains harbor at least one TPS system, the hrpA genes display significant C-terminal sequence variation. Meningococcal genes encoding the TPS effector proteins and their transporters are closely associated and transcribed into a single mRNA. HrpA proteins are translocated across the meningococcal outer membrane by their cognate transporters HrpB and mainly released into the environment. During this process, HrpA is proteolytically processed to a mature 180-kDa form. In contrast to other known TPS systems, immature HrpA proteins are stable in the absence of HrpB and accumulate within the bacterial cell. A small percentage of mature HrpA remains associated with the bacteria and contributes to the interaction of meningococci with epithelial cells.
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Blackwell, L. J., et J. A. Borowiec. « Human replication protein A binds single-stranded DNA in two distinct complexes ». Molecular and Cellular Biology 14, no 6 (juin 1994) : 3993–4001. http://dx.doi.org/10.1128/mcb.14.6.3993-4001.1994.
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Human replication protein A, a single-stranded DNA (ssDNA)-binding protein, is a required factor in eukaryotic DNA replication and DNA repair systems and has been suggested to function during DNA recombination. The protein is also a target of interaction for a variety of proteins that control replication, transcription, and cell growth. To understand the role of hRPA in these processes, we examined the binding of hRPA to defined ssDNA molecules. Employing gel shift assays that "titrated" the length of ssDNA, hRPA was found to form distinct multimeric complexes that could be detected by glutaraldehyde cross-linking. Within these complexes, monomers of hRPA utilized a minimum binding site size on ssDNA of 8 to 10 nucleotides (the hRPA8-10nt complex) and appeared to bind ssDNA cooperatively. Intriguingly, alteration of gel shift conditions revealed the formation of a second, distinctly different complex that bound ssDNA in roughly 30-nucleotide steps (the hRPA30nt complex), a complex similar to that described by Kim et al. (C. Kim, R. O. Snyder, and M. S. Wold, Mol. Cell. Biol. 12:3050-3059, 1992). Both the hRPA8-10nt and hRPA30nt complexes can coexist in solution. We speculate that the role of hRPA in DNA metabolism may be modulated through the ability of hRPA to bind ssDNA in these two modes.
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Blackwell, L. J., et J. A. Borowiec. « Human replication protein A binds single-stranded DNA in two distinct complexes. » Molecular and Cellular Biology 14, no 6 (juin 1994) : 3993–4001. http://dx.doi.org/10.1128/mcb.14.6.3993.
Texte intégralRésumé :
Human replication protein A, a single-stranded DNA (ssDNA)-binding protein, is a required factor in eukaryotic DNA replication and DNA repair systems and has been suggested to function during DNA recombination. The protein is also a target of interaction for a variety of proteins that control replication, transcription, and cell growth. To understand the role of hRPA in these processes, we examined the binding of hRPA to defined ssDNA molecules. Employing gel shift assays that "titrated" the length of ssDNA, hRPA was found to form distinct multimeric complexes that could be detected by glutaraldehyde cross-linking. Within these complexes, monomers of hRPA utilized a minimum binding site size on ssDNA of 8 to 10 nucleotides (the hRPA8-10nt complex) and appeared to bind ssDNA cooperatively. Intriguingly, alteration of gel shift conditions revealed the formation of a second, distinctly different complex that bound ssDNA in roughly 30-nucleotide steps (the hRPA30nt complex), a complex similar to that described by Kim et al. (C. Kim, R. O. Snyder, and M. S. Wold, Mol. Cell. Biol. 12:3050-3059, 1992). Both the hRPA8-10nt and hRPA30nt complexes can coexist in solution. We speculate that the role of hRPA in DNA metabolism may be modulated through the ability of hRPA to bind ssDNA in these two modes.
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Neil, R. Brock, et Michael A. Apicella. « Role of HrpA in Biofilm Formation of Neisseria meningitidis and Regulation of the hrpBAS Transcripts ». Infection and Immunity 77, no 6 (16 mars 2009) : 2285–93. http://dx.doi.org/10.1128/iai.01502-08.
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ABSTRACT Two-partner secretion systems of gram-negative organisms are utilized in adherence, invasion, and biofilm formation. The HrpAB proteins of Neisseria meningitidis are members of a two-partner secretion system, and HrpA is established as being important to adherence and intracellular escape. This study set out to determine the expression pattern of members of the hrpBAS putative operon and to find a functional role for the HrpA protein. The upregulation of these genes was found in situations of anaerobiosis and cell contact. These observations prompted the study of the function of HrpA in biofilms on human bronchial epithelial cells. HrpA mutants in encapsulated and unencapsulated NMB strains demonstrated biofilm growth equivalent to that of the wild-type strain at 6 h but a decreased ability to form biofilms at 48 h. Biofilms formed by hrpA mutants for 48 h on collagen-coated coverslips demonstrated significant reductions compared to those of wild-type strains. Taken together, these observations imply a role for HrpA in the biofilm structure. Further analysis demonstrated the presence of HrpA on the surface of the bacterium.
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Deng, Wen-Ling, Gail Preston, Alan Collmer, Chun-Jung Chang et Hsiou-Chen Huang. « Characterization of the hrpC and hrpRSOperons of Pseudomonas syringae Pathovars Syringae, Tomato, and Glycinea and Analysis of the Ability of hrpF,hrpG, hrcC, hrpT, and hrpVMutants To Elicit the Hypersensitive Response and Disease in Plants ». Journal of Bacteriology 180, no 17 (1998) : 4523–31. http://dx.doi.org/10.1128/jb.180.17.4523-4531.1998.
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The species Pseudomonas syringae encompasses plant pathogens with differing host specificities and corresponding pathovar designations. P. syringae requires the Hrp (type III protein secretion) system, encoded by a 25-kb cluster ofhrp and hrc genes, in order to elicit the hypersensitive response (HR) in nonhosts or to be pathogenic in hosts. DNA sequence analysis of the hrpC and hrpRSoperons of P. syringae pv. syringae 61 (brown spot of beans), P. syringae pv. glycinea U1 (bacterial blight of soybeans), and P. syringae pv. tomato DC3000 (bacterial speck of tomatos) revealed that the 13 genes comprising the right half of the hrp cluster (including those in the previously sequenced hrpZ operon) are conserved and identically arranged. The hrpC operon is comprised of hrpF,hrpG, hrcC, hrpT, and hrpV. hrcC encodes a putative outer membrane protein that is conserved in all type III secretion systems. The other four genes appear to be characteristic of group I Hrp systems, such as those possessed byP. syringae and Erwinia amylovora. The predicted products of these four genes in P. syringae pv. syringae 61 are HrpF (8 kDa), HrpG (15.4 kDa), HrpT (7.5 kDa), and HrpV (13.4 kDa). HrpT is a putative outer membrane lipoprotein. HrpF, HrpG, and HrpV are all hydrophilic proteins lacking N-terminal signal peptides. The HrpG, HrcC, HrpT, and HrpV proteins of P. syringae pathovars syringae and tomato (the two most divergent pathovars) had at least 76% amino acid identity with each other, whereas the HrpF proteins of these two pathovars had only 36% amino acid identity. The HrpF proteins of P. syringae pathovars syringae and glycinea also showed significant similarity to the HrpA pilin protein of P. syringae pathovar tomato. Functionally nonpolar mutations were introduced into each of the genes in thehrpC operon of P. syringae pv. syringae 61 by insertion of an nptII cartridge lacking a transcription terminator. The mutants were assayed for their ability to elicit the HR in nonhost tobacco leaves or to multiply and cause disease in host bean leaves. Mutations in hrpF, hrcC, andhrpT abolished or greatly reduced the ability of P. syringae pv. syringae 61 to elicit the HR in tobacco. ThehrpG mutant had only weakly reduced HR activity, and the activity of the hrpV mutant was indistinguishable from that of the wild type. Each of the mutations could be complemented, but surprisingly, the hrpV subclone caused a reduction in the HR elicitation ability of the ΔhrpV::nptIImutant. The hrpF and hrcC mutants caused no disease in beans, whereas the hrpG, hrpT, and hrpV mutants had reduced virulence. Similarly, thehrcC mutant grew little in beans, whereas the other mutants grew to intermediate levels in comparison with the wild type. These results indicate that HrpC and HrpF have essential functions in the Hrp system, that HrpG and HrpT contribute quantitatively but are not essential, and that HrpV is a candidate negative regulator of the Hrp system.
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Blackwell, L. J., J. A. Borowiec et I. A. Mastrangelo. « Single-stranded-DNA binding alters human replication protein A structure and facilitates interaction with DNA-dependent protein kinase. » Molecular and Cellular Biology 16, no 9 (septembre 1996) : 4798–807. http://dx.doi.org/10.1128/mcb.16.9.4798.
Texte intégralRésumé :
Human replication protein A (hRPA) is an essential single-stranded-DNA-binding protein that stimulates the activities of multiple DNA replication and repair proteins through physical interaction. To understand DNA binding and its role in hRPA heterologous interaction, we examined the physical structure of hRPA complexes with single-stranded DNA (ssDNA) by scanning transmission electron microscopy. Recent biochemical studies have shown that hRPA combines with ssDNA in at least two binding modes: by interacting with 8 to 10 nucleotides (hRPA8nt) and with 30 nucleotides (hRPA30nt). We find the relatively unstable hRPA8nt complex to be notably compact with many contacts between hRPA molecules. In contrast, on similar lengths of ssDNA, hRPA30nt complexes align along the DNA and make few intermolecular contacts. Surprisingly, the elongated hRPA30nt complex exists in either a contracted or an extended form that depends on ssDNA length. Therefore, homologous-protein interaction and available ssDNA length both contribute to the physical changes that occur in hRPA when it binds ssDNA. We used activated DNA-dependent protein kinase as a biochemical probe to detect alterations in conformation and demonstrated that formation of the extended hRPA30nt complex correlates with increased phosphorylation of the hRPA 29-kDa subunit. Our results indicate that hRPA binds ssDNA in a multistep pathway, inducing new hRPA alignments and conformations that can modulate the functional interaction of other factors with hRPA.
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Sangeeta et Arnab Bhattacherjee. « Interdomain dynamics in human Replication Protein A regulates kinetics and thermodynamics of its binding to ssDNA ». PLOS ONE 18, no 1 (19 janvier 2023) : e0278396. http://dx.doi.org/10.1371/journal.pone.0278396.
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Human Replication Protein A (hRPA) is a multidomain protein that interacts with ssDNA intermediates to provide the latter much-needed stability during DNA metabolism and maintain genomic integrity. Although the ssDNA organization with hRPA was studied recently through experimental means, characterizing the underlying mechanism at the atomic level remains challenging because of the dynamic domain architecture of hRPA and poorly understood heterogeneity of ssDNA-protein interactions. Here, we used a computational framework, precisely tailored to capture protein-ssDNA interactions, and investigated the binding of hRPA with a 60 nt ssDNA. Two distinct binding mechanisms are realized based on the hRPA domain flexibility. For a rigid domain architecture of hRPA, ssDNA binds sequentially with hRPA domains, resulting in slow association kinetics. The binding pathway involves the formation of stable and distinct intermediate states. On contrary, for a flexible domain architecture of hRPA, ssDNA binds synergistically to the A and B domains followed by the rest of hRPA. The domain dynamics in hRPA alleviates the free energy cost of domain orientation necessary for specific binding with ssDNA, leading to fast association kinetics along a downhill binding free energy landscape. An ensemble of free energetically degenerate intermediate states is encountered that makes it arduous to characterize them structurally. An excellent match between our results with the available experimental observations provides new insights into the rich dynamics of hRPA binding to ssDNA and in general paves the way to investigate intricate details of ssDNA-protein interactions, crucial for cellular functioning.
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Haapalainen, Minna, Kristin van Gestel, Minna Pirhonen et Suvi Taira. « Soluble Plant Cell Signals Induce the Expression of the Type III Secretion System of Pseudomonas syringae and Upregulate the Production of Pilus Protein HrpA ». Molecular Plant-Microbe Interactions® 22, no 3 (mars 2009) : 282–90. http://dx.doi.org/10.1094/mpmi-22-3-0282.
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Type III protein secretion is essential for the pathogenicity of Pseudomonas syringae on its host plants. Expression of HrpA, a major component of the type III secretion system (T3SS)-associated pilus, was studied both in plant leaves and in vitro using reporter genes. We found that induction of the hrpA promoter was stronger in plants than in vitro, and that the induction was enhanced by both host and nonhost plants of P. syringae pv. tomato. In vitro, the expression was enhanced by cell-free exudates from plant cell suspension cultures, added into the minimal medium. Further analysis of the plant-cell-derived, hrpA-inducing factors showed that they were small and water-soluble compounds, which could signal P. syringae the proximity of living plant cells. We also studied the production and secretion of native HrpA protein in vitro, and detected a plant-signal-dependent increase in HrpA secretion. In contrast to HrpA, the intracellular accumulation or secretion of the other T3SS-dependent proteins were not significantly increased, despite the presence of plant cell-derived, promoter-inducing factors. Thus, the accumulation of HrpA pilin seems to be subjected to a distinct post-transcriptional regulation.
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Grass, Lena M., Jan Wollenhaupt, Tatjana Barthel, Iwan Parfentev, Henning Urlaub, Bernhard Loll, Eberhard Klauck, Haike Antelmann et Markus C. Wahl. « Large-scale ratcheting in a bacterial DEAH/RHA-type RNA helicase that modulates antibiotics susceptibility ». Proceedings of the National Academy of Sciences 118, no 30 (21 juillet 2021) : e2100370118. http://dx.doi.org/10.1073/pnas.2100370118.
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Many bacteria harbor RNA-dependent nucleoside-triphosphatases of the DEAH/RHA family, whose molecular mechanisms and cellular functions are poorly understood. Here, we show that the Escherichia coli DEAH/RHA protein, HrpA, is an ATP-dependent 3 to 5′ RNA helicase and that the RNA helicase activity of HrpA influences bacterial survival under antibiotics treatment. Limited proteolysis, crystal structure analysis, and functional assays showed that HrpA contains an N-terminal DEAH/RHA helicase cassette preceded by a unique N-terminal domain and followed by a large C-terminal region that modulates the helicase activity. Structures of an expanded HrpA helicase cassette in the apo and RNA-bound states in combination with cross-linking/mass spectrometry revealed ratchet-like domain movements upon RNA engagement, much more pronounced than hitherto observed in related eukaryotic DEAH/RHA enzymes. Structure-based functional analyses delineated transient interdomain contact sites that support substrate loading and unwinding, suggesting that similar conformational changes support RNA translocation. Consistently, modeling studies showed that analogous dynamic intramolecular contacts are not possible in the related but helicase-inactive RNA-dependent nucleoside-triphosphatase, HrpB. Our results indicate that HrpA may be an interesting target to interfere with bacterial tolerance toward certain antibiotics and suggest possible interfering strategies.
Thèses sur le sujet "HrpA protein"
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Alegria, Marcos Castanheira. « Identificação de interações proteína-proteína envolvendo os produtos dos Loci hrp, vir e rpf do fitopatógeno Xanthomonas axonopodis pv. citri ». Universidade de São Paulo, 2004. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-22082016-162740/.
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O Cancro Cítrico, um dos mais graves problemas fitossanitários da citricultura atual, é uma doença causada pelo fitopatógeno Xanthomonas axonopodis pv. citri (Xac). Um estudo funcional do genoma de Xac foi iniciado com o intuito de identificar interações proteína-proteína envolvidas em processos de patogenicidade de Xac. Através da utilização do sistema duplo-híbrido de levedura, baseado nos domínios de ligação ao DNA e ativação da transcrição do GAL4, nós analisamos os principais componentes dos mecanismos de patogenicidade de Xac, incluindo o Sistema de Secreção do Tipo III (TTSS), Sistema de Secreção do Tipo IV (TFSS) e Sistema de \"Quorum Sensing\" composto pelas proteínas Rpf. Componentes desses sistemas foram utilizados como iscas na triagem de uma biblioteca genômica de Xac. O TTSS é codificado pelos genes denominados hrp (\"hypersensitive response and pathogenicity\"), hrc (\"hrp conserved\") e hpa (\"hrp associated\") localizados no locus hrp do cromossomo de Xac. Esse sistema de secreção é capaz de translocar proteínas efetoras do citoplasma bacteriano para o interior da célula hospedeira. Nossos resultados mostraram novas interações proteínaproteína entre componentes do próprio TTSS além de associações específicas com uma proteína hipotética: 1) HrpG, um regulador de resposta de um sistema de dois componentes responsável pela expressão dos genes hrp, e XAC0095, uma proteína hipotética encontrada apenas em Xanthomonas spp; 2) HpaA, uma proteína secretada pelo TTSS, HpaB e o domínio C-terminal da HrcV; 3) HrpB1, HrpD6 e HrpW, 4) HrpB2 e HrcU e 5) interações homotrópicas envolvendo a ATPase HrcN. Em Xac, foram encontrados dois loci vir que codificam proteínas que possuem similaridade com componentes do TFSS envolvido em processos de conjugação/secreção bacteriana: TFSS-plasmídeo localizado no plasmídeo pXAC64 e TFSS-cromossomo localizado no cromossomo de Xac. O TFSS-plasmídeo, o qual possui maior similaridade com sistemas de conjugação, mostrou interações envolvendo proteínas cujos genes estão localizados na mesma região do plasmídeo pXAC64: 1) interação homotrópica da TrwA; 2) XACb0032 e XACb0033; 3) interações homotrópicas da proteína XACb0035; 4) VirB1 e VirB9; 5) XACb0042 e VirB6; 6) XACb0043 e XACb0021b. O TFSS-cromossomo apresentou interações envolvendo as proteínas: 1) VirD4 e um grupo de 12 proteínas que contém similaridade entre si, incluindo XAC2609 cujo gene encontra-se no locus vir, 2) XAC2609 e XAC2610; 3) Interações homotrópicas da VirB11; 4) XAC2622 e VirB9. A análise do sistema de \"Quorum-Sensing\" composto pelas proteínas Rpf mostrou interações envolvendo componentes do próprio sistema: 1) RpfC e RpfF; 2) RpfC e RpfG; 3) interações homotrópicas da RpfF; 4) RpfC e CmfA, uma proteína similar a Cmf de Dictyostelium discoideum que, neste organismo, é fundamental para processos de \"quorum-sensing\". As interações proteína-proteína encontradas permitiram-nos entender melhor a composição, organização e regulação dos fatores envolvidos na patogenicidade de Xac.Citrus Canker, caused by the bacterial plant pathogen Xanthomonas axonopodis pv. citri (Xac) presents one of the most serious problems to Brazilian citriculture. We have initiated a project to identify protein-protein interactions involved in pathogenicity of Xac. Using a yeast two-hybrid system based on GAL4 DNA-binding and activation domains, we have focused on identifying interactions involving subunits, regulators and substrates of: Type Three Secretion System (TTSS), Type Four Secretion System (TFSS) and Quorum Sensing/Rpf System. Components of these systems were used as baits to screening a random Xac genomic library. The TTSS is coded by the hrp (hypersensitive response and pathogenicity), hrc (hrp conserved) and hpa (hrp associated) genes in the chromosomal hrp locus. This secretion system can translocate efector proteins from the bacterial cytoplasm into the host cells. We have identified several previously uncharacterized interactions involving: 1) HrpG, a two-component system response regulator responsible for the expression of Xac hrp operons, and XAC0095, a previously uncharacterized protein encountered only in Xanthomonas spp; 2) HpaA, a protein secreted by the TTSS, HpaB and the C-terminal domain HrcV; 3) HrpB1, HrpD6 and HrpW; 4) HrpB2 and HrcU; 5) Homotropic interactions were also identified for the ATPase HrcN. Xac contains two virB gene clusters, one on the chromosome and one on the pXAC64 plasmid, each of which codes for a unique and previously uncharacterized TFSS. Components of the TFSS of pXAC64, which is most similar to conjugation systems, showed interactions involving proteins coded by the same locus: 1) Homotropic interactions of TrwA; 2) XACb0032 and XACb0033; 3) XAC0035 homotropic interactions; 4) VirB1 and VirB9; 5) XACb0042 and VirB6; 6) XACb0043 and XACb0021 b. Components of the chromosomal TFSS exhibited interactions involving: 1) VirD4 and a group of 12 uncharacterized proteins with a common C-terminal domain motif, include XAC2609 whose gene resides within the vir locus; 2) XAC2609 and XAC261 O; 3) Homotropic interactions of VirB11; 4) XAC2622 and VirB9. Analysis of Quorum Sensing/Rpf System components revealed interactions between the principal Rpf proteins which control Xanthomonas quorum sensing: 1) RpfC and RpfF; 2) RpfC and RpfG; 3) RpfF homotropic interactions; 4) RpfC and CmfA, a protein that presents similarity with Cmf (conditioned medium factor) of Dictyostelium discoideum, which contrais quorum sensing in this organism. The protein-protein interactions that we have detected reveal insights into the composition, organization and regulation of these important mechanisms involved in Xanthomonas pathogenicity.
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Winck, Flavia Vischi. « Estudo das proteinas HrpF e AvrXacE2 na patogenicidade de Xanthomonas axonopodis pv. citri ». [s.n.], 2007. http://repositorio.unicamp.br/jspui/handle/REPOSIP/314772.
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Orientador: Marcos Antonio MachadoDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A bactéria Xanthomonas axonopodis pv. citri (Xac) é o agente causador do cancro cítrico, doença que leva a severas perdas econômicas devido à contaminação e à erradicação de plantas de citros. Com o seqüenciamento completo do genoma de Xac, vários genes supostamente envolvidos com a patogenicidade de Xac foram identificados. Os genes ligados à resposta de hipersensibilidade (hrp) e avirulência (avr) em geral estão relacionados à patogenicidade de Xanthomonas, entretanto, poucos estudos funcionais destes genes de Xac foram feitos. Foram construídas linhagens mutantes de Xac para a perda de função dos genes hrpF e avrXacE2 e, nas análises in vivo, foi verificado que hrpF está envolvido na patogenicidade de Xac e é essencial para a manifestação dos sintomas primários da doença. A mutação de avrXacE2 não provocou alterações na capacidade de Xac em provocar os sintomas do cancro, portanto, este gene não parece ser essencial para a patogenicidade da bactéria, podendo não estar envolvido diretamente na patogenicidade de Xac. Os genes hrpF e avrXacE2 foram clonados em vetores de expressão e foram realizados testes de indução da expressão destas proteínas em sistemas heterólogos. Somente a proteína AvrXacE2 foi expressa, purificada e submetida a teste de interação com as proteínas citoplasmáticas da linhagem mutante de Xac para o gene avrXacE2. Os testes de interações não confirmaram a identificação de proteínas com afinidade específica pela proteína recombinante AvrXacE2. A proteína HrpF não foi super-expressa em sistema heterólogo. Nas análises de proteoma comparativo da linhagem de Xac selvagem versus linhagem mutante para o gene hrpF, foram detectadas alterações na expressão de proteínas citoplasmáticas e "pericelulares". Com base nas observações pode-se supor que HrpF possa influenciar processos celulares relacionados à respostas à situações de estresse e não somente atuar na translocação de moléculas efetoras via T3SS. A partir do que foi exposto neste trabalho, sugere-se que as técnicas de estudos funcionais de genes e análises proteômicas podem conjuntamente permitir que novos mecanismos relacionados a patogenicidade de Xac sejam interpretados. Com os mutantes produzidos neste estudo, espera-se criar condições para novos ensaios funcionais visando a melhor compreensão da patogenicidade de Xac e buscar novas formas de combate ao cancro cítrico
Abstract: The bacterium Xanthomonas axonopodis pv. citri (Xac) is the causative agent of the citrus canker disease, which leads to economic losses due the contamination and erradication of citrus plants. The complete sequencing of its genome identified a number of genes supposedly involved with pathogenicity. Genes that code for hipersensitivity response (hrp) and avirulence (avr), in general, are related to the pathogenicity of Xanthomonas, however, only a few functional studies of these genes in Xac have been made. Here we report findings based on genomics and proteomics methods for Xac. Mutant strains of Xac for genes hrpF and avrXacE2 and in vivo assays demonstrated that hrpF is strongly involved in the pathogenicity of Xac and is essential for the manifestation of the primary symptoms of the citrus canker. On the other hand, the lack of avrXacE2 expression did not result in modifications in the capacity of Xac to elicite the symptoms of canker, therefore, this gene does not seem to be essential for the pathogenicity of the bacterium. The genes hrpF and avrXacE2 were cloned in expression vectors and tests of induction of the expression of these proteins in heterologous systems were carried out. The protein AvrXacE2 was expressed, purified and tested on interaction assays with cytoplasmic proteins of the mutant of Xac for the gene avrXacE2. The tests of interactions had not confirmed the identification of proteins with specific affinity for the recombinant protein AvrXacE2. The protein HrpF was not overexpressed in heterologous system. In the comparative proteome of the wild versus mutant strains for hrpF, modifications in the cytoplasmic protein expression and "pericellular" expression levels were detected. We postulate that, besides acting as a translocator of molecules through T3SS, HrpF may influence stress-related cellular responses. Thus, it is an opportune time to highlight the new and different ways in which HrpF serves Xac function. Moreover, we can assume that the techniques of functional genomics and proteomics analyses will clarify the mechanisms of pathogenicity used by Xac to cause citrus canker and, thus, enable the search for additional information to control the disease
Mestrado
Bioquimica
Mestre em Biologia Funcional e Molecular
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Al-Fartusie, Falah Sumoon Daghal. « Engineering and characterisation of novel protein covalent linkages in horseradish peroxidase (HRP) : effect on structure and function ». Thesis, University of Sussex, 2011. http://sro.sussex.ac.uk/id/eprint/7422/.
Texte intégralRésumé :
Both mammalian and bacterial peroxidases contain novel covalent linkages. In the former a sulfonium linkage to a critical Met residue is thought to modify the normal planarity of the haem affecting the functional properties. Following on from the work of Metcalfe et al., 2004 which showed that such links could be engineered in ascorbate peroxidase, horseradish peroxidase (HRP) has been used as convenient model system to try and understand the structural and electronic effects of engineered covalent linkages. Previous work in the group (Cali, 2008) had shown that mutation of Ser167 to Met in HRP-C* resulted in autocatalytic cross-linking on incubation with hydrogen peroxide. In this thesis, two additional HRP variants, S167Y and S167W were studied and a new novel structural linkage discovered. The UV/Vis spectrum of the S167Y variant suggested a more 6-coordinate high spin character. The molar extinction coefficients were markedly increased, 180 mM-1 cm-1 for S167Y and 135 mM-1 cm-1 for S167W, compared to 100 mM-1 cm-1 for the WT enzyme, consistent with a more 6 coordinate high spin character normally seen in lignin peroxidase. In contrast, the dissociation constant (Kd) of the S167W variant mutant for the aromatic donor BHA was hardly affected, whilst that of the S167Y variant increased two-fold relative to the WT, implying a significant perturbation of the aromatic donor binding site and / or the associated haem-linked hydrogen bonding network. After peroxide treatment the haem group of the S167Y variant could not be extracted into acid butanone in contrast to the WT. Only a proportion of the haem could be extracted even from the untreated S167Y variant, implying that a substantial fraction of the protein had formed the haem-protein linkage during folding and purification. These results were confirmed during reverse phase HPLC and MALDI-TOF / ESI mass spectroscopy measurements. The haem and protein completely co-eluted in the case of peroxide treated S167Y, while only ~50% of the haem was linked to the protein in the untreated as isolated enzyme. The MALDI-TOF and ESI mass spectrum showed that there was a large increase (614 Da) in the mass of the linked S167Y protein, compared to that of the unlinked enzyme. Unlike the sulfonium linkage obtained earlier, treatment with hydrogen peroxide was unnecessary to observe this increase. Interestingly, the 100% unlinked S167Y protein could only be isolated if enzyme was prepared in the presence of an efficient peroxidase substrate as an antioxidant scavenger. It appears that a Tyr residue at position 167 is highly reactive with respect of the haem vinyl side chain forming a spontaneous covalent link not otherwise seen in nature. Pre steady-state comparison of the intermediates has shown that Compound I was formed essentially normally at near WT rates, however its stability was greatly affected in the S167Y variant (linked or unlinked), the life time being decreased to ~0.04 s, compared to of the WT enzyme, where it was ~80 s. The substrate preference of the cross-linked S167Y variant was also altered. Stopped-flow measurements of the individual rate constants for the partial reactions of the catalytic cycle with luminol as reducing substrate revealed an increase in the rate of reduction of Compound I to Compound II (k2). The X-ray crystal structure of S167Y variant was solved to 1.7 Å resolution and the structure has been modelled and determined by x-ray crystallography. The x-ray structure reveals an unanticipated linkage containing an additional ring structure bonded to the engineered Tyr. ESI mass measurements supported this structure.
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Perez, Humberto Rodriguez. « HrcA de Caulobacter crescentus e Xylella fastidiosa : estudos comparativos de seqüências e desenvolvimento de modelo estrutural ». Universidade de São Paulo, 2002. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-27092018-143815/.
Texte intégralRésumé :
O gene hrcA é encontrado em quase todos os ramos da árvore filogenética das eubactérias, e seu produto, a proteína HrcA, funciona como repressor da expressão dos operons de choque térmico groESL e dnaKJ, ligando-se à seqüência repetida invertida denominada CIRCE (controlling inverted repeat of chaperonin expression) presente na região regulatória destes operons. O sistema HrcA-CIRCE está, portanto, amplamente representado nas eubactérias. Particularmente, em Caulobacter crescentus, uma α-proteobactéria, este sistema está envolvido no controle da expressão do operon groESL durante o ciclo celular da bactéria. Conhecer a estrutura e as interações de HrcA é importante para entender este processo. Neste trabalho são apresentadas as análises de seqüência das HrcA\'s de C. crescentus e de Xylella fastidiosa, uma proteobactéria do grupo γ, as quais são muito similares. Este estudo levou à proposta de um modelo estrutural com a delimitação dos domínios da proteína, os dobramentos de cada domínio, com base nas interações da HrcA de C. crescentus com o elemento CIRCE e ATP, que estão sendo caracterizadas em nosso laboratório, assim como a atribuição de aminoácidos e motivos conservados funcionais. Adicionalmente, embora a expressão da HrcA recombinante de X. fastidiosa não tenha tido sucesso, a HrcA recombinante de C. crescentus purificada tem se prestado aos ensaios espectroscópicos, ainda que tenha sido detectada uma microagregação que está sendo enfrentada com um protocolo de purificação baseado no uso de α ciclodextrina. Os estudos espectroscópicos preliminares da HrcA C. crescentus dão suporte ao modelo estrutural proposto.The hrcA gene is found in almost all branches of the filogenetic tree of eubacteria, and its product, the protein HrcA, functions as a repressor regulating the expression of the heat shock operons groESL and dnaKJ, by binding to the inverted repeat sequence called CIRCE (controlling inverted repeat of chaperonin expression). The system HrcA-CIRCE, therefore, is widely represented in eubacteria. Specifically in Caulobacter crescentus, an α-proteobacterium, this system is involved in the cell-cycle control of groESL expression (Baldini et al, 1998). Knowledge of the structure of HrcA and its interactions is important to understand this process. This work presents the analysis of the sequences of HrcA from C. crescentus and Xylella fastidiosa, a proteobacterium of the γ group, which are very similar. A structural model has been proposed, with protein domain delimitation, specific domain folding, based on known interactions of C. crescentus HrcA with the CIRCE element and ATP, obtained in our laboratory, as well as assignment of functional residues and conserved motifs. Additionally, even though no sucess was obtained the expression of recombinant HrcA from X. fastidiosa, purified recombinant HrcA from C. crescentus has been shown to be suitable for spectroscopic studies, in spite of microagregation observed, which is being faced with a purification protocol based on the use of α cyclodextrin. The preliminary spectroscopic studies of HrcA from C. crescentus support the proposed structural model.
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Nadhom, Hama. « Protein Microparticles for Printable Bioelectronics ». Thesis, Linköpings universitet, Biosensorer och bioelektronik, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-119637.
Texte intégralRésumé :
In biosensors, printing involves the transfer of materials, proteins or cells to a substrate. It offers many capabilities thatcan be utilized in many applications, including rapid deposition and patterning of proteins or other biomolecules.However, issues such as stability when using biomaterials are very common. Using proteins, enzymes, as biomaterialink require immobilizations and modifications due to changing in the structural conformation of the enzymes, whichleads to changes in the properties of the enzyme such as enzymatic activity, during the printing procedures andrequirements such as solvent solutions. In this project, an innovative approach for the fabrication of proteinmicroparticles based on cross-linking interchange reaction is presented to increase the stability in different solvents.The idea is to decrease the contact area between the enzymes and the surrounding environment and also preventconformation changes by using protein microparticles as an immobilization technique for the enzymes. The theory isbased on using a cross-linking reagent trigging the formation of intermolecular bonds between adjacent proteinmolecules leading to assembly of protein molecules within a CaCO3 template into a microparticle structure. TheCaCO3 template is removed by changing the solution pH to 5.0, leaving behind pure highly homogenous proteinmicroparticles with a size of 2.4 ± 0.2 μm, according to SEM images, regardless of the incubation solvents. Theenzyme model used is Horse Radish Peroxidase (HRP) with Bovine Serum Albumin (BSA) and Glutaraldehyde (GL)as a cross-linking reagent. Furthermore, a comparison between the enzymatic activity of the free HRP and the BSAHRPprotein microparticles in buffer and different solvents are obtained using Michaelis-Menten Kinetics bymeasuring the absorption of the blue product produced by the enzyme-substrate interaction using a multichannelspectrophotometer with a wavelength of 355 nm. 3,3’,5,5’-tetramethylbenzidine (TMB) was used as substrate. As aresult, the free HRP show an enzymatic activity variation up to ± 50 % after the incubation in the different solventswhile the protein microparticles show much less variation which indicate a stability improvement. Moreover, printingthe microparticles require high microparticle concentration due to contact area decreasing. However, usingmicroparticles as a bioink material prevent leakage/diffusion problem that occurs when using free protein instead.
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Cali, Khasim Cumar. « Towards the Design of New Functional Properties in Horseradish peroxidase (HRP) : Engineering a Covalent link between the Haem and the Protein ». Thesis, University of Sussex, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.506842.
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Susin, Michelle Fernanda. « Análise funcional das proteínas HrcA, GroES/GroEL e DnaK/DnaJ em Caulobacter crescentus ». Universidade de São Paulo, 2005. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-14062016-171416/.
Texte intégralRésumé :
O operon groESL de C. crescentus apresenta dupla regulação. A indução deste operon por choque térmico é dependente do fator sigma de choque térmico σ32. A temperaturas fisiológicas, a expressão de groESL apresenta regulação temporal durante o ciclo celular da bactéria e o controle envolve a proteína repressora HrcA e o elemento CIRCE (controlling inverted repeat of chaperonin expression). Para estudar a atividade da proteína repressora in vitro, produzimos e purificamos de E. coli a HrcA de C. creseentus contendo uma cauda de histidinas e a ligação especifica ao elemento CIRCE foi analisada em ensaios de migração retardada em gel de poliacrilamida (EMRGP). A quantidade de DNA retardada pela ligação a HrcA aumentou significativamente na presença de GroES/GroEL, sugerindo que estas proteínas modulam a atividade de HrcA. Corroboração desta modulação foi obtida analisando fusões de transcrição da região regulatória de groESL com o gene lacZ, em células de C. crescentus produzindo diferentes quantidades de GroES/EL. HrcA contendo as substituições Pro81 AJa e Arg87Ala, aminoácidos que se localizam no domínio putativo de ligação ao DNA da proteína, mostraram ser deficientes na ligação a CIRCE, tanto in vitro como in vivo. Em adição, HrcA Ser56Ala expressa na mesma célula juntamente com a proteína selvagem produziu um fenótipo dominante-negativo, indicando que a HrcA de C. crescentus liga-se a CIRCE como um oligômero, provavelmente um dímero. As tentativas de obtenção de mutantes nulos para os genes groESL ou dnaKJ falharam, indicando que as proteínas GroES/GroEL e DnaK/DnaJ são essenciais em C. crescentus, mesmo a temperaturas normais. Foram então construídas no laboratório as linhagens mutantes condicionais SG300 e SG400 de C. crescentus, onde a expressão de groESL e de dnaKJ, respectivamente, está sob controle de um promotor induzido por xilose (PxyIX). Estas linhagens foram caracterizadas quanto á sua morfologia em condições permissivas ou restritivas, assim como quanto à capacidade de sobrevivência frente a vários tipos de estresse. As células da linhagem SG300, exauridas de GroES/GroEL, são resistentes ao choque térmico a 42°C e são capazes de adquirir alguma termotolerância. Entretanto, estas células são sensíveis aos estresses oxidativo, salino e osmótico. As células da linhagem SG400, exauridas de DnaKlJ, são sensíveis ao choque térmico, à exposição a etanol e ao congelamento, e são incapazes de adquirir termotolerância. Além disso, tanto as células exauridas de GroES/GroEL quanto as exauridas de DnaK/DnaJ apresentam problemas na sua morfologia. As células de SG300 exauridas de GroES/GroEL formam filamentos longos que possuem constrições fundas e irregulares. As células de SG400 exauridas de DnaK/DnaJ são apenas um pouco mais alongadas que as células pré-divisionais selvagens e a maioria das células não possuem septo. Estas observações indicam bloqueio da divisão celular, que deve ocorrer em diferentes estágios em cada linhagem.In Caulobacter crescentus, the groESL operon presents a dual type of control. Heat shock induction of the operon is dependent on the heat shock sigma factor σ-32. At physiological temperatures, groESL expression is cell cycle regulated and the control involves the repressor protein HrcA and the element CIRCE (controlling inverted repeat of chaperonin ~xpression). To study the activity of HrcA in vitro, we produced and purified from E. coli a histidine-tagged version of the protein, and specific binding to the CIRCE element was analyzed in electrophoretic mobility shift assays (EMSA). The amount of retarded DNA increased significantly in the presence of GroES/GroEL, suggesting that these proteins modulate HrcA activity. Further evidence of this modulation was obtained using lacZ transcription fusions with the groESL regulatory region in C. crescentus cells producing different amounts of GroES/GroEL. The mutants proteins HrcA Pro81Ala and HrcA Arg87Ala, that contain amino acid substitutions in the putative DNA-bindíng domain of the protein, were found to be deficient in binding to CIRCE in vitro and in vivo. Furthermore, HrcA Ser56Ala expressed together with the wild type protein within the same cell, produced a dominant-negative phenotype, indicating that C. crescentus HrcA binds to CIRCE in an oligomeric form, most likely as a dimer. Attempts to obtain null mutants for groESL or dnaKJ were unsuccessful indicating the importance of GroES/GroEL and DnaK/lDnaJ to the survival of C. crescentus cells. Conditional mutants were then constructed in our laboratory in which groESL and dnaKJ expression is under the control ofaxylose inducible promoter (PxyIX) , giving rise to strains SG300 and SG400, respectively. These strains were characterized in regard to their morphology under permissive and restrictive conditions, as well as their viability under different types of environmental stresses. SG300 cells depleted of GroES/GroEL are resistant to heat shock at 42°C and can acquire some thermotolerance, but they are sensitive to oxidative, saline and osmotic stresses. SG400 cells depleted of DnaKlJ are quite sensitive to heat shock, ethanol and freezing, and are unable of acquiring thermotolerance. Cells depleted of either GroES/EL or DnaKlJ also present morphological problems. SG300 cells depleted of GroES/EL form long and pinched filaments. SG400 cells depleted of DnaKlJ are only somewhat more elongated than wild-type predivisional cells and most cells do not present septum. These observations indicate a cell division arrest, which should occur at different stages in each strain.
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Willie, Nigani. « Plasmodium falciparum Histidine-rich Protein 2 Gene Variation and Malaria Detection in Madagascar and Papua New Guinea ». Case Western Reserve University School of Graduate Studies / OhioLINK, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=case1519326080906088.
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Guo, Yu-Wun, et 郭昱彣. « Characterization of HrpY and HrpW proteins in Acidovorax avenae subsp. avenae CH12 ». Thesis, 2013. http://ndltd.ncl.edu.tw/handle/30213248148798845526.
Texte intégralRésumé :
碩士國立中興大學
生物科技學研究所
101
Acidovorax avenae subsp. avenae (Aaa) CH12 causes bacterial leaf stripe disease on corn. The bacterial proteins secreted via type III secretion system (T3SS) which is encoded by a hrp/hrc cluster are the major virulence factors to cause diseases in host plants or elicit the hypersensitive response (HR) on nonhost plants. The genes residing the region between hrcT and GALA genes in hrp/hrc cluster of A. avenae isolated from different host are variable. To elucidate whether the diversity is involved in virulence or not, the two ORFs annotated in this region from AaaCH12 were characterized in this study. These two ORFs were named as hrpY and hrpW based on their amino acid sequence shared 99% and 98% identities with those from AaaN1141, respectively. Moreover, HrpY and HrpW from AaaN1141 strain were predicted to be harpin proteins, which are glycine rich and thermal stable, could be an HR elicitor, have no N-terminal signal peptide and secreted via T3SS. In this study, HrpY and HrpW proteins from AaaCH12 are glycine rich and have no N-terminal signal peptide. The crude extracted or purified HrpY-His6 and HrpW-His6 proteins which were overexpressed by T7 RNA polymerase system could elicit the HR on Nicotiana tabacum with or without heat treatment, suggesting that HrpY-His6 and HrpW-His6 proteins are thermal stable. A pectate lyase domain residing in C-terminus of HrpW from AaaCH12 has pectate lyase activity based on the pectate lyase assay using polyglacturonic acid as a substrate. Gel filtration assay showed both purified HrpY-His6 and HrpW-His6 could be detected as about 2000 kDa in size. HrpY-His6 protein was applied to raise a polyclonal antibody in rabbit for Western blot analysis. Under T3SS inducing condition by cultured in modified XVM2 medium, both HrpY and HrpW could be secreted in wild type but not in T3SS-deficient hrcV mutant of AaaCH12, suggesting that HrpY and HrpW are secreted via T3SS. The results strongly suggest that HrpY and HrpW are harpin-like proteins. In addition, hrpY, hrpW and hrpYW deletion mutants were generated by using unmarked gene deletion mutagenesis. The hrpY, hrpW and hrpYW mutants elicited the delay HR on N. tabacum. The hrpY and hrpYW deletion mutants also reduced the virulence of Aaa CH12 by decreasing disease lesion length and bacterial growth in corn leaves, but the hrpW deletion mutant did not. Additionally, the hrpYW double mutant is significantly less virulent than hrpY, suggesting that HrpYW proteins have an additive effect in virulence on corns.
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BIANCALANI, CAROLA. « Anti-infective environmentally friendly molecules against plant pathogenic Gram-negative bacteria ». Doctoral thesis, 2017. http://hdl.handle.net/2158/1087786.
Texte intégralRésumé :
The market globalisation and the climate change are contributing substantially to the possible and rapid spread of alien and invasive plant pathogens in areas where they were previously absent, or are intensifying the incidence and severity of endemic pathogens, thus contributing significantly to increase the possible threats to the agricultural sector. Moreover, the lack of effective alternative molecules to copper compounds in plant protection, whose negative effects on both human health and environmental protection, have been neglected for far too long, and the need to adapt to European legislation, have led the operators in plant protection sector to reflect about the urgent need to implement a cultural revolution in which major innovation efforts are required.
The study was carried out in order to achieve the following main aims: I) analysing pathogenic and virulence systems of phytopathogenic Gram-negative bacteria such as the Type Three Secretion System (TTSS), and in particular the main structural protein of TTSS pilus, i.e. “HrpA”, in order to design molecules able to block the pathogenicity and virulence of these bacteria without undermining their viability; II) verifying the in vitro and in vivo efficacy of anti-infective molecules, such as small oligopeptides and polyphenolic extracts obtained in a circular economy framework, to reduce or to block symptoms development caused by plant pathogenic bacteria; finally, as a future objective to analyse a possible correlation among virulence systems, fitness and efflux pumps related to xenobiotic compounds extrusion in phytopathogenic bacteria, in order to identify underdeveloped targets, against which innovative molecules can be designed.
Chapitres de livres sur le sujet "HrpA protein"
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Bozsó, Zoltán, Péter G. Ott et Zoltán Klement. « Hr-Positive Phenotype of the Pseudomonas syringae pv. syringae hrpK Mutant and hrp Gene Superinduction in Tobacco Leaves Treated with Protein Synthesis Inhibitors ». Dans Developments in Plant Pathology, 122–26. Dordrecht : Springer Netherlands, 1997. http://dx.doi.org/10.1007/978-94-011-5472-7_22.
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Hutcheson, Steven W. « The hrp Cluster of Pseudomonas syringae : a Pathogenicity Island Encoding a Type III Protein Translocation Complex ? » Dans Pathogenicity Islands and Other Mobile Virulence Elements, 309–29. Washington, DC, USA : ASM Press, 2014. http://dx.doi.org/10.1128/9781555818173.ch16.
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Peñaloza-Vázquez, A., G. M. Preston, A. C. Collmer et C. L. Bender. « The Hrp Protein Secretion System is not Required for Coronatine Biosynthesis in Pseudomonas syringae pv. tomato DC3000 ». Dans Plant Pathogenic Bacteria, 205–8. Dordrecht : Springer Netherlands, 2001. http://dx.doi.org/10.1007/978-94-010-0003-1_46.
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Collmer, Alan, B. H. Kvitko, J. E. Morello, K. R. Munkvold, H. S. Oh et C. F. Wei. « Exploring the Functions of Proteins Secreted by the Hrp Type III Secretion System of Pseudomonas syringae ». Dans Pseudomonas syringae Pathovars and Related Pathogens – Identification, Epidemiology and Genomics, 229–37. Dordrecht : Springer Netherlands, 2008. http://dx.doi.org/10.1007/978-1-4020-6901-7_24.
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Van Gijsegem, Frédérique, Eliane Farcy, Matthieu Arlat, Claudine Zischek, Clare Gough, Stéphane Genin, Marc Marenda, Samantha Vernhettes et Christian Boucher. « Role of Proteins Encoded by the Pseudomonas Solanacearum Hrp Regulon in the Control of Plant-Bacteria Interactions ». Dans Advances in Molecular Genetics of Plant-Microbe Interactions, 65–69. Dordrecht : Springer Netherlands, 1994. http://dx.doi.org/10.1007/978-94-011-0177-6_10.
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Hutcheson, Steven W. « The hrp-Encoded Protein Export Systems of Pseudomonas syringae and Other Plant Pathogenic Bacteria and Their Role in Pathogenicity ». Dans Plant-Microbe Interactions, 145–79. Boston, MA : Springer US, 1997. http://dx.doi.org/10.1007/978-1-4615-6019-7_7.
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Van Gijsegem, F., M. Marenda, B. Brito, J. Vasse, C. Zischek, S. Genin, M. Guéneron, P. Barberis, M. Arlat et C. Boucher. « The Ralstonia solanacearum hrp Gene Region : Role of the Encoded Proteins in Interactions with Plants and Regulation of Gene Expression ». Dans Bacterial Wilt Disease, 178–83. Berlin, Heidelberg : Springer Berlin Heidelberg, 1998. http://dx.doi.org/10.1007/978-3-662-03592-4_26.
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Shawa, Remmy, Fons Coomans, Helen Cox et Leslie London. « Access to Effective Diagnosis and Treatment for Drug-Resistant Tuberculosis : Deepening the Human Rights-Based Approach ». Dans Ethics and Drug Resistance : Collective Responsibility for Global Public Health, 155–69. Cham : Springer International Publishing, 2020. http://dx.doi.org/10.1007/978-3-030-27874-8_10.
Texte intégralRésumé :
Abstract The lack of access to effective diagnosis and treatment for drug-resistant tuberculosis (DR-TB) remains a persistent ethical, human rights and public health challenge globally. In addressing this challenge, arguments based on a Human Rights-Based Approach (HRBA) to health have most often been focused on the Right to Health. However, a key challenge in multidrug-resistant (MDR-) and extensively drug-resistant (XDR-) TB is the glaring absence of scientific research; ranging from basic science and drug discovery through to implementation science once new tools have been developed. Although the Right to Enjoy the Benefits of Scientific Progress and its Applications (REBSP) is a little theorised human right, it has the potential to enrich our understanding and use of the Rights-Based Approach to health. In this chapter, we argue that States’ duties to respect, protect and fulfil the REBSP within and outside their borders is an important vehicle that can be drawn on to redress the lack of research into new drug development and appropriate use of existing drugs for DR-TB in high burden settings. We call for urgent attention to minimum core obligations for the REBSP and the need for a General Comment by a UN human rights monitoring body to provide for its interpretation. We also note that conceptualization of the REBSP has the potential to complement Right to Health claims intended to enhance access to treatment for DR-TB on a global scale.
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Holland, James, et Julian Webb. « 10. ‘Bringing Rights Home’ : Legal Method and the Convention Rights ». Dans Learning Legal Rules, 326–54. Oxford University Press, 2019. http://dx.doi.org/10.1093/he/9780198799900.003.0010.
Texte intégralRésumé :
In the twenty-first century, two important pan-European forces to which English law has been subject are the European Convention on Human Rights (ECHR) and the Human Rights Act (HRA) 1998. This chapter discusses the following: the scope, outline, and enforcement of the ECHR to identify and protect fundamental human rights and freedoms and the balancing of these freedoms against the sovereignty of Parliament; its incorporation into the HRA 1998; incorporation under the devolution Acts; the consequences for legal method; and practical and conceptual issues raised by the HRA 1998 around legal research and argumentation. It closes by looking at the prospects of a ‘British Bill of Rights’.
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Holland, James, et Julian Webb. « 9. ‘Bringing Rights Home’ : Legal Method and the Convention Rights ». Dans Learning Legal Rules, 304–34. Oxford University Press, 2022. http://dx.doi.org/10.1093/he/9780192849090.003.0009.
Texte intégralRésumé :
In the twenty-first century, two important pan-European forces to which English law has been subject are the European Convention on Human Rights (ECHR) and the Human Rights Act (HRA) 1998. This chapter discusses the following: the scope, outline, and enforcement of the ECHR to identify and protect fundamental human rights and freedoms and the balancing of these freedoms against the sovereignty of Parliament; its incorporation into the HRA 1998; incorporation under the devolution Acts; the consequences for legal method; and practical and conceptual issues raised by the HRA 1998 around legal research and argumentation. It closes by looking at the prospects of a ‘British Bill of Rights’.
Actes de conférences sur le sujet "HrpA protein"
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Perossi, Isabela Fernanda Spinelli, Mylena Mitie Saito, Giovanna Rossi Varallo, Jucimara Colombo et Débora Aparecida Pires de Campos Zuccari. « ANALYSIS OF OVERALL SURVIVAL IN BITCHES WITH BREAST CANCER USING TARGET PROTEINS RELATED TO THE PI3K/AKT/MTOR PATHWAY ». Dans Abstracts from the Brazilian Breast Cancer Symposium - BBCS 2021. Mastology, 2021. http://dx.doi.org/10.29289/259453942021v31s2007.
Texte intégralRésumé :
This study aims to verify survival in female dogs with breast tumor by analyzing the expression of target proteins PIK3CA, ZEB1, and ZEB2 belonging to the PI3K/AKT/mTOR pathway through immunohistochemistry (IHC) test in a retrospective study. The samples were obtained from dogs with breast cancer, previously identified by standard histopathological analysis, from which tissue microarray (TMA) blocks were made, and then immunohistochemical analyzes (IHC) were carried out using the development kit “REVEAL Polyvalent HRP-DAB Detection System,” for the proteins previously mentioned. For the purpose of prognostic analysis, these dogs were monitored for 540 days after surgical resection and survival was related to protein expression using the histoscore (HS) method. The HS is a measure to convert the IHC into quantitative values, and it is based on the intensity of the staining and the percentage of stained cells, ranging from 0 to 300. Individually through the analysis of the IHC, it was observed in the PIK3CA protein that from the HS = 164 the survival was on average 189 days, for ZEB1, the HS = 100 the survival was on average 438 days, and on the protein ZEB2 with the HS = 157, the survival was on average 178 days. Thus, the high expression of PIK3CA and ZEB2 proteins was correlated with lower survival in the dogs. In all studied proteins, it was observed that HS > 100 was correlated with a significant reduction in overall survival (p>0.05). The lower survival in female dogs with breast cancer after surgical resection was related to low rates of expression of PIK3CA and ZEB2 and, therefore, these can be considered as prognostic markers reserved for breast cancer in female dogs.
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Yezdimer, Eric, Nina Rittereiser, Ceren Yüce, Berthold Köhler et Matthias Reihmann. « Compatibility of Hydroxyproline Rich, Natural Proteins (HRPs) and Surfactants in Hard Surface Cleaner Formulations ». Dans Virtual 2021 AOCS Annual Meeting & Expo. American Oil Chemists’ Society (AOCS), 2021. http://dx.doi.org/10.21748/am21.389.
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Polat, Burak, et Mihai A. Diaconeasa. « On the Use of Probabilistic Risk Assessment for the Protection of Small Modular Reactors Against Terrorist Attacks ». Dans ASME 2021 International Mechanical Engineering Congress and Exposition. American Society of Mechanical Engineers, 2021. http://dx.doi.org/10.1115/imece2021-71504.
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Abstract Safety and security are two of the most important requirements of the nuclear industry. In the event of a potential problem, the consequences can have serious implications for the public and the environment. Measures should be taken against various hazards and threats by analyzing possible realistic scenarios. Therefore, probabilistic risk assessment is one of the necessary technologies to achieving safe and secure nuclear facilities. In the study, a limited scope probabilistic risk assessment was made for a possible terrorist attack against a generic small modular reactor (SMR). A possible attack threat was selected to develop scenarios by following a probabilistic risk assessment approach. In the scenarios created, terrorists have to pass all physical barriers that security guards protect. Thus, the decisions and actions of the security guards directly affect the result of the attack. To analyze these events, a human reliability assessment (HRA) was employed. In the first study, each security guard’s decision-making process was analyzed using the Standardized Plant Analysis Risk Human Reliability Assessment (SPAR-H) method. The purpose of its use in this study is to verify the SPAR-H method’s applicability for security applications. In this paper, we give the likelihoods of each security guard making a decision and taking action to prevent terrorists from passing obtained using the SPAR-H method. Besides, event tree and fault tree analyses were performed using the SAPHIRE PRA software. Finally, since the current HRA methods were designed for control room operators, we introduce a new model-based HRA methodology applicable for security guards to be used in physical security PRAs.
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Gonzalez, Rachel M., Li Ding, Ming Xiao et Xiaoping Zhang. « Abstract 3193 : A new approach for colocalization of proteins using HRP and AP enzyme detection in paraffin embedded tissues ». Dans Proceedings : AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012 ; Chicago, IL. American Association for Cancer Research, 2012. http://dx.doi.org/10.1158/1538-7445.am2012-3193.
Texte intégralRapports d'organisations sur le sujet "HrpA protein"
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Coplin, David, Isaac Barash et Shulamit Manulis. Role of Proteins Secreted by the Hrp-Pathways of Erwinia stewartii and E. herbicola pv. gypsophilae in Eliciting Water-Soaking Symptoms and Initiating Galls. United States Department of Agriculture, juin 2001. http://dx.doi.org/10.32747/2001.7580675.bard.
Texte intégralRésumé :
Many bacterial pathogens of plants can inject pathogenicity proteins into host cells using a specialized type III secretion system encoded by hrpgenes. This system deliver effector proteins, into plant cells that function in both susceptible and resistant interactions. We have found that the virulence of Erwinia stewartii(Es; syn. Pantoea stewartii) and Erwinia herbicola pv. gypsophilae (Ehg, syn. Pantoea agglomerans), which cause Stewart's wilt of corn and galls on Gypsophila, respectively, depends on hrpgenes. The major objectives of this project were: To increase expression of hrpgenes in order to identify secreted proteins; to identify genes for proteins secreted by the type-III systems and determine if they are required for pathogenicity; and to determine if the secreted proteins can function within eukaryotic cells. We found that transcription of the hrp and effector genes in Es and Ehg is controlled by at least four genes that constitute a regulatory cascade. Environmental and/or physiological signaling appears to be mediated by the HrpX/HrpY two component system, with HrpX functioning as a sensor-kinase and HrpY as a response regulator. HrpYupregulateshrpS, which encodes a transcriptional enhancer. HrpS then activates hrpL, which encodes an alternate sigma factor that recognizes "hrp boxes". All of the regulatory genes are essential for pathogenicity, except HrpX, which appears only to be required for induction of the HR in tobacco by Es. In elucidating this regulatory pathway in both species, we made a number of significant new discoveries. HrpX is unusual for a sensor-kinase because it is cytoplasmic and contains PAS domains, which may sense the redox state of the bacterium. In Es, a novel methyl-accepting protein may function upstream of hrpY and repress hrp gene expression in planta. The esaIR quorum sensing system in Es represses hrp gene expression in Es in response to cell-density. We have discovered six new type III effector proteins in these species, one of which (DspE in Ehg and WtsE in Es) is common to both pathogens. In addition, Es wtsG, which is a homolog of an avrPpiB from P. syringae pv. pisi, and an Ehg ORF, which is a homolog of P. syringae pv. phaseolicola AvrPphD, were both demonstrated to encode virulence proteins. Two plasmidborne, Ehg Hop proteins, HsvG and PthG, are required for infection of gypsophilia, but interestingly, PthG also acts as an Avr elicitor in beets. Using a calmodulin-dependent adenylate cyclase (cyaA) reporter gene, we were successful in demonstrating that an HsvG-CyaA fusion protein can be transferred into human HeLa cells by the type-III system of enteropathogenic E. coli. This is a highly significant accomplishment because it is the first direct demonstration that an effector protein from a plant pathogenic bacterium is capable of being translocated into a eukaryotic cell by a type-III secretion system. Ehg is considered a limiting factor in Gypsophila production in Israel and Stewart’s Wilt is a serious disease in the Eastern and North Central USA, especially on sweet corn in epidemic years. We believe that our basic research on the characterization of type III virulence effectors should enable future identification of their receptors in plant cells. This may lead to novel approaches for genetically engineering resistant plants by modifying their receptors or inactivating effectors and thus blocking the induction of the susceptible response. Alternatively, hrp gene regulation might also provide a target for plant produced compounds that interfere with recognition of the host by the pathogen. Such strategies would be broadly applicable to a wide range of serious bacterial diseases on many crops throughout the USA and Israel.
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Coplin, David L., Shulamit Manulis et Isaac Barash. roles Hrp-dependent effector proteins and hrp gene regulation as determinants of virulence and host-specificity in Erwinia stewartii and E. herbicola pvs. gypsophilae and betae. United States Department of Agriculture, juin 2005. http://dx.doi.org/10.32747/2005.7587216.bard.
Texte intégralRésumé :
Gram-negative plant pathogenic bacteria employ specialized type-III secretion systems (TTSS) to deliver an arsenal of pathogenicity proteins directly into host cells. These secretion systems are encoded by hrp genes (for hypersensitive response and pathogenicity) and the effector proteins by so-called dsp or avr genes. The functions of effectors are to enable bacterial multiplication by damaging host cells and/or by blocking host defenses. We characterized essential hrp gene clusters in the Stewart's Wilt of maize pathogen, Pantoea stewartii subsp. stewartii (Pnss; formerly Erwinia stewartii) and the gall-forming bacterium, Pantoea agglomerans (formerly Erwinia herbicola) pvs. gypsophilae (Pag) and betae (Pab). We proposed that the virulence and host specificity of these pathogens is a function of a) the perception of specific host signals resulting in bacterial hrp gene expression and b) the action of specialized signal proteins (i.e. Hrp effectors) delivered into the plant cell. The specific objectives of the proposal were: 1) How is the expression of the hrp and effector genes regulated in response to host cell contact and the apoplastic environment? 2) What additional effector proteins are involved in pathogenicity? 3) Do the presently known Pantoea effector proteins enter host cells? 4) What host proteins interact with these effectors? We characterized the components of the hrp regulatory cascade (HrpXY ->7 HrpS ->7 HrpL ->7 hrp promoters), showed that they are conserved in both Pnss and Fag, and discovered that the regulation of the hrpS promoter (hrpSp) may be a key point in integrating apoplastic signals. We also analyzed the promoters recognized by HrpL and demonstrated the relationship between their composition and efficiency. Moreover, we showed that promoter strength can influence disease expression. In Pnss, we found that the HrpXY two-component signal system may sense the metabolic status of the bacterium and is required for full hrp gene expression in planta. In both species, acyl-homoserine lactone-mediated quorum sensing may also regulate epiphytic fitness and/or pathogenicity. A common Hrp effector protein, DspE/WtsE, is conserved and required for virulence of both species. When introduced into corn cells, Pnss WtsE protein caused water-soaked lesions. In other plants, it either caused cell death or acted as an Avr determinant. Using a yeast- two-hybrid system, WtsE was shown to interact with a number of maize signal transduction proteins that are likely to have roles in either programmed cell death or disease resistance. In Pag and Pab, we have characterized the effector proteins HsvG, HsvB and PthG. HsvG and HsvB are homologous proteins that determine host specificity of Pag and Pab on gypsophila and beet, respectively. Both possess a transcriptional activation domain that functions in yeast. PthG was found to act as an Avr determinant on multiple beet species, but was required for virulence on gypsophila. In addition, we demonstrated that PthG acts within the host cell. Additional effector genes have been characterized on the pathogenicity plasmid, pPATHₚₐg, in Pag. A screen for HrpL- regulated genes in Pnsspointed up 18 candidate effector proteins and four of these were required for full virulence. It is now well established that the virulence of Gram-negative plant pathogenic bacteria is governed by Hrp-dependent effector proteins. However; the mode of action of many effectors is still unresolved. This BARD supported research will significantly contribute to the understanding of how Hrp effectors operate in Pantoea spp. and how they control host specificity and affect symptom production. This may lead to novel approaches for genetically engineering plants resistant to a wide range of bacterial pathogens by inactivating the Hrp effectors with "plantabodies" or modifying their receptors, thereby blocking the induction of the susceptible response. Alternatively, innovative technologies could be used to interfere with the Hrp regulatory cascade by blocking a critical step or mimicking plant or quorum sensing signals.