scholarly journals Xanthomonas oryzae pv. oryzae XKK.12 Contains an AroQγ Chorismate Mutase That Is Involved in Rice Virulence

2010 ◽  
Vol 100 (3) ◽  
pp. 262-270 ◽  
Author(s):  
Giuliano Degrassi ◽  
Giulia Devescovi ◽  
Joseph Bigirimana ◽  
Vittorio Venturi
Keyword(s):  
Pathogenic Bacteria ◽  
Shikimate Pathway ◽  
Aromatic Amino Acids ◽  
Xanthomonas Oryzae ◽  
Chorismate Mutase ◽  
In Planta ◽  
Knock Out ◽  
Key Enzyme ◽  
Precise Role

Chorismate mutase (CM) is a key enzyme in the shikimate pathway which is responsible for the synthesis of aromatic amino acids. There are two classes of CMs, AroQ and AroH, and several pathogenic bacteria have been reported to possess a subgroup of CMs designated AroQγ. These CMs are usually exported to the periplasm or outside the cell; in a few cases, they have been reported to be involved in virulence and their precise role is currently unknown. Here, we report that the important rice pathogen Xanthomonas oryzae pv. oryzae XKK.12 produces an AroQγ CM which we have purified and characterized from spent supernatants. This enzyme is synthesized in planta and X. oryzae pv. oryzae knock-out mutants are hypervirulent to rice. The role of this enzyme in X. oryzae pv. oryzae rice virulence is discussed.

scholarly journals Genetic and biochemical identification of the chorismate mutase from Corynebacterium glutamicum

Microbiology ◽  
2009 ◽  
Vol 155 (10) ◽  
pp. 3382-3391 ◽  
Author(s):  
Pan-Pan Li ◽  
Ya-Jun Liu ◽  
Shuang-Jiang Liu
Keyword(s):  
Corynebacterium Glutamicum ◽  
Shikimate Pathway ◽  
Hypothetical Protein ◽  
Chorismate Mutase ◽  
Cloning And Expression ◽  
Physical Interaction ◽  
Structural Class ◽  
Important Amino Acid ◽  
Key Enzyme ◽  
Biochemical Identification

Chorismate mutase (CM) catalyses the rearrangement of chorismate to prephenate and is also the first and the key enzyme that diverges the shikimate pathway to either tryptophan (Trp) or phenylalanine (Phe) and tyrosine (Tyr). Corynebacterium glutamicum is one of the most important amino acid producers for the fermentation industry and has been widely investigated. However, the gene(s) encoding CM has not been experimentally identified in C. glutamicum. In this study, the ncgl0819 gene, which was annotated as ‘conserved hypothetical protein’ in the C. glutamicum genome, was genetically characterized to be essential for growth in minimal medium, and a mutant deleted of ncgl0819 was a Phe and Tyr auxotroph. Genetic cloning and expression of ncgl0819 in Escherichia coli resulted in the formation of a new protein (NCgl0819) having CM activity. It was concluded that ncgl0819 encoded the CM of C. glutamicum (CM0819). CM0819 was demonstrated to be a homodimer and is a new member of the monofunctional CMs of the AroQ structural class. The CM0819 activity was not affected by Phe, Tyr or Trp. Two 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthases (DS0950 and DS2098, formerly NCgl0950 and NCgl2098) had been previously identified from C. glutamicum. CM0819 significantly stimulated DAHP synthase (DS2098) activity. Physical interaction between CM0819 and DS2098 was observed. When CM0819 was present, DS2098 activity was subject to allosteric inhibition by chorismate and prephenate. Conserved hypothetical proteins homologous to CM0819 were identified in all known Corynebacterium genomes, suggesting a universal occurrence of CM0819-like CMs in the genus Corynebacterium.

scholarly journals In Planta Colonization and Role of T6SS in Two Rice Kosakonia Endophytes

2020 ◽  
Vol 33 (2) ◽  
pp. 349-363 ◽  
Author(s):  
Susan Mosquito ◽  
Iris Bertani ◽  
Danilo Licastro ◽  
Stéphane Compant ◽  
Michael P. Myers ◽  
...  
Keyword(s):  
Comparative Genomics ◽  
Growth Promotion ◽  
Rice Seed ◽  
In Planta ◽  
Root Cortex ◽  
Knock Out ◽  
Colonization Ability ◽  
Secretory System ◽  
Secretome Profiling

Endophytes live inside plants and are often beneficial. Kosakonia is a novel bacterial genus that includes many diazotrophic plant-associated isolates. Plant–bacteria studies on two rice endophytic Kosakonia beneficial strains were performed, including comparative genomics, secretome profiling, in planta tests, and a field release trial. The strains are efficient rhizoplane and root endosphere colonizers and localized in the root cortex. Secretomics revealed 144 putative secreted proteins, including type VI secretory system (T6SS) proteins. A Kosakonia T6SS genomic knock-out mutant showed a significant decrease in rhizoplane and endosphere colonization ability. A field trial using rice seed inoculated with Kosakonia spp. showed no effect on plant growth promotion upon nitrogen stress and microbiome studies revealed that Kosakonia spp. were significantly more present in the inoculated rice. Comparative genomics indicated that several protein domains were enriched in plant-associated Kosakonia spp. This study highlights that Kosakonia is an important, recently classified genus involved in plant–bacteria interaction.

scholarly journals The role of swnR gene on the biosynthesis pathway of the swainsonine in Metarhizium anisopliae

2020 ◽  
Author(s):  
Lu Sun ◽  
Runjie Song ◽  
Jinglong Wang ◽  
Yu Zhang ◽  
Yanli Zhu ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Metarhizium Anisopliae ◽  
Type Strain ◽  
Fermentation Broth ◽  
Biosynthesis Pathway ◽  
Mutant Type ◽  
Knock Out ◽  
Q Exactive ◽  
Precise Role

Abstract Background: Swainsonine (SW) is the principal toxic ingredient of locoweeds, and is produced by fungi including Metarhizium anisopliae, Slafractonia leguminicola, and Alternaria oxytropis (found in locoweeds of Oxytropis). Studies of the SW biosynthesis pathway in these fungi have demonstrated the requirement for a swnK gene and the presence of a variety of other SWN cluster genes, but have not determined a precise role for the swnR gene, which encodes a NADB Rossmann-fold reductase, nor if it is necessary for the biosynthesis of SW. In this study, we used homologous recombination (HR) to knock out the swnR gene of M. anisopliae to determine its effect on the SW biosynthesis pathway.Results: The concentration of SW was measured in the fermentation broth of M. anisopliae at 1 d, 3 d, 5 d and 7 d using a Q Exactive Mass Spectrometer. The gene for swnR was detected by RT-qPCR. To determine the role of the swnR gene in the SW biosynthesis pathway of M. anisopliae, we used PEG-mediated homologous recombination (HR) to transform a wild-type strain (WT) with a Benomyl (ben)-resistant fragment to knock out the swnR gene producing a mutant-type strain (MT). A complemented-type (CT) strain was produced by adding a complementation vector that contains the glufosinate (herbicide) resistance (bar) gene as a marker. The content of SW decreased, but was not eliminated in the fermentation broth of the MT strain, and returned to the original level in the CT strain.Conclusions: These results indicate that the swnR gene plays a crucial role in the SW biosynthesis pathway of M. anisopliae, but suggests that another gene in the fungus may share the function of swnR.

Regulation and pathophysiological implications of UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE) as the key enzyme of sialic acid biosynthesis

2009 ◽  
Vol 390 (7) ◽  
Author(s):  
Stefan O. Reinke ◽  
Gerhard Lehmer ◽  
Stephan Hinderlich ◽  
Werner Reutter
Keyword(s):  
Sialic Acid ◽  
Crucial Role ◽  
Feedback Inhibition ◽  
Acid Biosynthesis ◽  
Knock Out ◽  
Acetylneuraminic Acid ◽  
Kinase C ◽  
Key Enzyme ◽  
Hereditary Inclusion Body Myopathy

AbstractThe key enzyme for the biosynthesis ofN-acetylneuraminic acid, from which all other sialic acids are formed, is the bifunctional enzyme UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE). GNE is a highly conserved protein found throughout the animal kingdom. Its highest expression is seen in the liver and placenta. GNE is regulated by a variety of biochemical means, including tetramerization promoted by the substrate UDP-GlcNAc, phosphorylation by protein kinase C and feedback inhibition by CMP-Neu5Ac, which is defect in the human disease sialuria. GNE knock-out in mice leads to embryonic lethality, emphasizing the crucial role of this key enzyme for sialic acid biosynthesis. The metabolic capacity to synthesize sialic acid and CMP-sialic acid upon ManNAc loads is amazingly high. An additional characteristic of GNE is its interaction with proteins involved in the regulation of development, which might play a crucial role in the hereditary inclusion body myopathy. Due to the importance of increased concentrations of tumor-surface sialic acid, first attempts to find inhibitors of GNE have been successful.

scholarly journals XadM, a Novel Adhesin of Xanthomonas oryzae pv. oryzae, Exhibits Similarity to Rhs Family Proteins and Is Required for Optimum Attachment, Biofilm Formation, and Virulence

2012 ◽  
Vol 25 (9) ◽  
pp. 1157-1170 ◽  
Author(s):  
Binod B. Pradhan ◽  
Manish Ranjan ◽  
Subhadeep Chatterjee
Keyword(s):  
Pathogenic Bacteria ◽  
Surface Protein ◽  
Growth Conditions ◽  
Xanthomonas Oryzae ◽  
Carbohydrate Binding ◽  
Significant Similarity ◽  
Reading Frame ◽  
Family Protein ◽  
Bacterial Blight Pathogen

By screening a transposon-induced mutant library of Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, we have identified a novel 5.241-kb open reading frame (ORF) named xadM that is required for optimum virulence and colonization. This ORF encodes a protein, XadM, of 1,746 amino acids that exhibits significant similarity to Rhs family proteins. The XadM protein contains several repeat domains similar to a wall-associated surface protein of Bacillus subtilis, which has been proposed to be involved in carbohydrate binding. The role of XadM in X. oryzae pv. oryzae adhesion was demonstrated by the impaired ability of an xadM mutant strain to attach and form biofilms. Furthermore, we show that XadM is exposed on the cell surface and its expression is regulated by growth conditions and plays an important role in the early attachment and entry inside rice leaves. Interestingly, XadM homologs are present in several diverse bacteria, including many Xanthomonas spp. and animal-pathogenic bacteria belonging to Burkholderia spp. This is the first report of a role for XadM, an Rhs family protein, in adhesion and virulence of any pathogenic bacteria.

scholarly journals Metabolic dependency of chorismate in Plasmodium falciparum suggests an alternative source for the ubiquinone biosynthesis precursor

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Ana Lisa Valenciano ◽  
Maria L. Fernández-Murga ◽  
Emilio F. Merino ◽  
Nicole R. Holderman ◽  
Grant J. Butschek ◽  
...  
Keyword(s):  
Malaria Parasite ◽  
Shikimate Pathway ◽  
Aromatic Amino Acids ◽  
Defined Medium ◽  
Current Evidence ◽  
Head Group ◽  
Main Role ◽  
Alternative Source ◽  
Folate Biosynthesis

Abstract The shikimate pathway, a metabolic pathway absent in humans, is responsible for the production of chorismate, a branch point metabolite. In the malaria parasite, chorismate is postulated to be a direct precursor in the synthesis of p-aminobenzoic acid (folate biosynthesis), p-hydroxybenzoic acid (ubiquinone biosynthesis), menaquinone, and aromatic amino acids. While the potential value of the shikimate pathway as a drug target is debatable, the metabolic dependency of chorismate in P. falciparum remains unclear. Current evidence suggests that the main role of chorismate is folate biosynthesis despite ubiquinone biosynthesis being active and essential in the malaria parasite. Our goal in the present work was to expand our knowledge of the ubiquinone head group biosynthesis and its potential metabolic dependency on chorismate in P. falciparum. We systematically assessed the development of both asexual and sexual stages of P. falciparum in a defined medium in the absence of an exogenous supply of chorismate end-products and present biochemical evidence suggesting that the benzoquinone ring of ubiquinones in this parasite may be synthesized through a yet unidentified route.

scholarly journals Role of the FeoB Protein and Siderophore in Promoting Virulence of Xanthomonas oryzae pv. oryzae on Rice

2010 ◽  
Vol 192 (12) ◽  
pp. 3187-3203 ◽  
Author(s):  
Alok Pandey ◽  
Ramesh V. Sonti
Keyword(s):  
Gene Cluster ◽  
Functional Characterization ◽  
Biosynthetic Gene Cluster ◽  
Xanthomonas Oryzae ◽  
In Planta ◽  
Growth Deficiency ◽  
Constitutive Production ◽  
Serious Disease

ABSTRACT Xanthomonas oryzae pv. oryzae causes bacterial blight, a serious disease of rice. Our analysis revealed that the X. oryzae pv. oryzae genome encodes genes responsible for iron uptake through FeoB (homolog of the major bacterial ferrous iron transporter) and a siderophore. A mutation in the X. oryzae pv. oryzae feoB gene causes severe virulence deficiency, growth deficiency in iron-limiting medium, and constitutive production of a siderophore. We identified an iron regulated xss gene cluster, in which xssABCDE ( X anthomonas siderophore synthesis) and xsuA ( X anthomonas siderophore utilization) genes encode proteins involved in biosynthesis and utilization of X. oryzae pv. oryzae siderophore. Mutations in the xssA, xssB, and xssE genes cause siderophore deficiency and growth restriction under iron-limiting conditions but are virulence proficient. An xsuA mutant displayed impairment in utilization of native siderophore, suggesting that XsuA acts as a specific receptor for a ferric-siderophore complex. Histochemical and fluorimetric assays with gusA fusions indicate that, during in planta growth, the feoB gene is expressed and that the xss operon is not expressed. This study represents the first report describing a role for feoB in virulence of any plant-pathogenic bacterium and the first functional characterization of a siderophore-biosynthetic gene cluster in any xanthomonad.

scholarly journals Classification of the glyphosate target enzyme (5-enolpyruvylshikimate-3-phosphate synthase)

2020 ◽  
Author(s):  
Lyydia Leino ◽  
Tuomas Tall ◽  
Marjo Helander ◽  
Irma Saloniemi ◽  
Kari Saikkonen ◽  
...  
Keyword(s):  
Gut Microbiome ◽  
Shikimate Pathway ◽  
Aromatic Amino Acids ◽  
Research Field ◽  
Human Gut ◽  
Human Gut Microbiome ◽  
The Core ◽  
Key Enzyme ◽  
Phosphate Synthase

ABSTRACTGlyphosate is the most common broad-spectrum herbicide. It targets the key enzyme of the shikimate pathway, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), which synthesizes three essential aromatic amino acids (phenylalanine, tyrosine and tryptophan) in plants. Because the shikimate pathway is also found in many prokaryotes and fungi, the widespread use of glyphosate may have unsuspected impacts on the diversity and composition of microbial communities, including the human gut microbiome. Here, we introduce the first bioinformatics method to assess the potential sensitivity of organisms to glyphosate based on the type of EPSPS enzyme. We have precomputed a dataset of EPSPS sequences from thousands of species that will be an invaluable resource to advancing the research field. This novel methodology can classify sequences from >90% of eukaryotes and >80% of prokaryotes. A conservative estimate from our results shows that 54% of species in the core human gut microbiome are sensitive to glyphosate.

scholarly journals The role of swnR gene on the biosynthesis pathway of the swainsonine in Metarhizium anisopliae

2019 ◽  
Author(s):  
Lu Sun ◽  
Runjie Song ◽  
Jinglong Wang ◽  
Yiling Liu ◽  
Yu Zhang ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Metarhizium Anisopliae ◽  
Type Strain ◽  
Fermentation Broth ◽  
Biosynthesis Pathway ◽  
Mutant Type ◽  
Knock Out ◽  
Q Exactive ◽  
Precise Role

AbstractSwainsonine (SW) is the principal toxic ingredient of locoweeds, and is produced by fungi including Metarhizium anisopliae, Slafractonia leguminicola, and Alternaria oxytropis. Studies of the SW biosynthesis pathway in these fungi have demonstrated the requirement for a swnK gene and the presence of a variety of other SWN cluster genes, but have not determined a precise role for the swnR gene, which encodes a NADB Rossmann-fold reductase, nor if it is necessary for the biosynthesis of SW. In this study, we used homologous recombination (HR) to knock out the swnR gene of M. anisopliae to determine its effect on the SW biosynthesis pathway. The concentration of SW was measured in the fermentation broth of M. anisopliae at 1 d, 3 d, 5 d and 7 d using a Q Exactive Mass Spectrometer. The gene for swnR was detected by RT-qPCR. To determine the role of the swnR gene in the SW biosynthesis pathway of M. anisopliae, we used PEG-mediated homologous recombination (HR) to transform a wild-type strain (WT) with a Benomyl (ben)-resistant fragment to knock out the swnR gene producing a mutant-type strain (MT). A complemented-type (CT) strain was produced by adding a complementation vector that contains the glufosinate (herbicide) resistance (bar) gene as a marker. The content of SW decreased, but was not eliminated in the fermentation broth of the MT strain, and returned to the original level in the CT strain. These results indicate that the swnR gene plays a crucial role in the SW biosynthesis pathway of M. anisopliae, but suggests that another gene in the fungus may share the function of swnR.

scholarly journals Cloning and Characterization of an Esophageal-Gland-Specific Chorismate Mutase from the Phytoparasitic Nematode Meloidogyne javanica

1999 ◽  
Vol 12 (4) ◽  
pp. 328-336 ◽  
Author(s):  
Kris N. Lambert ◽  
Keith D. Allen ◽  
Ian M. Sussex
Keyword(s):  
Cell Formation ◽  
Shikimate Pathway ◽  
Aromatic Amino Acids ◽  
Meloidogyne Javanica ◽  
Giant Cells ◽  
Chorismate Mutase ◽  
Root Knot Nematode ◽  
Esophageal Gland ◽  
Giant Cell Formation

Root-knot nematodes are obligate plant parasites that alter plant cell growth and development by inducing the formation of giant feeder cells. It is thought that nematodes inject secretions from their esophageal glands into plant cells while feeding, and that these secretions cause giant cell formation. To elucidate the mechanisms underlying the formation of giant cells, a strategy was developed to clone esophageal gland genes from the root-knot nematode Meloidogyne javanica. One clone, shown to be expressed in the nematode's esophageal gland, codes for a potentially secreted chorismate mutase (CM). CM is a key branch-point regulatory enzyme in the shikimate pathway and converts chorismate to prephenate, a precursor of phenylalanine and tyrosine. The shikimate pathway is not found in animals, but in plants, where it produces aromatic amino acids and derivative compounds that play critical roles in growth and defense. Therefore, we hypothesize that this CM is involved in allowing nematodes to parasitize plants.

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