scholarly journals N-Deacetylation of Sinorhizobium meliloti Nod Factors Increases Their Stability in the Medicago sativa Rhizosphere and Decreases Their Biological Activity

2000 ◽  
Vol 13 (1) ◽  
pp. 72-79 ◽  
Author(s):  
Christian Staehelin ◽  
Michael Schultze ◽  
Ken Tokuyasu ◽  
Véréna Poinsot ◽  
Jean-Claude Promé ◽  
...  
Keyword(s):  
Biological Activity ◽  
Medicago Sativa ◽  
Sinorhizobium Meliloti ◽  
Colletotrichum Lindemuthianum ◽  
Signal Molecules ◽  
Glycosyl Hydrolases ◽  
Nod Factors ◽  
Nod Factor ◽  
Chemical Structures

Nod factors excreted by rhizobia are signal molecules that consist of a chitin oligomer backbone linked with a fatty acid at the nonreducing end. Modifications of the Nod factor structures influence their stability in the rhizosphere and their biological activity. To test the function of N-acetyl groups in Nod factors, NodSm-IV(C16:2,S) from Sinorhizobium meliloti was enzymatically N-deacetylated in vitro with purified chitin deacetylase from Colletotrichum lindemuthianum. A family of partially and completely deacetylated derivatives was produced and purified. The most abundant chemical structures identified by mass spectrometry were GlcN(C16:2)-GlcNAc-GlcNH2-GlcNAc(OH)(S), GlcN(C16:2)-GlcNAc-GlcNH2-GlcNH2(OH)(S), and GlcN(C16:2)-GlcNH2-GlcNH2-GlcNH2(OH)(S). In contrast to NodSm-IV(C16:2,S), the purified N-deacetylated derivatives were stable in the rhizosphere of Medicago sativa, indicating that the N-acetyl groups make the carbohydrate moiety of Nod factors accessible for glycosyl hydrolases of the host plant. The N-deacetylated derivatives displayed only a low level of activity in inducing root hair deformation. Furthermore, the N-deacetylated molecules were not able to stimulate Nod factor degradation by M. sativa roots, a response elicited by active Nod factors. These data show that N-acetyl groups of Nod factors are required for biological activity.

scholarly journals Activity of Sinorhizobium meliloti NodAB and NodH Enzymes on Thiochitooligosaccharides

2002 ◽  
Vol 184 (14) ◽  
pp. 4039-4043 ◽  
Author(s):  
Audrey M. Southwick ◽  
Lai-Xi Wang ◽  
Sharon R. Long ◽  
Yuan C. Lee
Keyword(s):  
Sinorhizobium Meliloti ◽  
Signal Molecules ◽  
Gene Products ◽  
Nod Factors ◽  
Nod Factor ◽  
Glycosidic Bonds

ABSTRACT Rhizobium bacteria synthesize signal molecules called Nod factors that elicit responses in the legume root during nodulation. Nod factors, modified N-acylated β-(1,4)-N-acetylglucosamine, are synthesized by the nodulation (nod) gene products. We tested the ability of three Sinorhizobium meliloti nod gene products to modify Nod factor analogs with thio linkages instead of O-glycosidic bonds in the oligosaccharide backbone.

scholarly journals Nod Factors of Rhizobium leguminosarum bv. viciae and Their Fucosylated Derivatives Stimulate a Nod Factor Cleaving Activity in Pea Roots and Are Hydrolyzed In Vitro by Plant Chitinases at Different Rates

2000 ◽  
Vol 13 (8) ◽  
pp. 799-807 ◽  
Author(s):  
Alexandra O. Ovtsyna ◽  
Michael Schultze ◽  
Igor A. Tikhonovich ◽  
Herman P. Spaink ◽  
Éva Kondorosi ◽  
...  
Keyword(s):  
Biological Activity ◽  
Sinorhizobium Meliloti ◽  
Rhizobium Leguminosarum ◽  
Nodule Development ◽  
Nod Factors ◽  
Degradation Rates ◽  
Pea Roots ◽  
The Stability ◽  
Plant Chitinases

Nod factors (NFs) are rhizobial lipo-chitooligosaccharide signals that trigger root nodule development in legumes. Modifications of NF structures influence their biological activity and affect their degradation by plant chitinases. Nodulation of certain pea cultivars by Rhizobium leguminosarum bv. viciae requires modification of NFs at the reducing end by either an O-acetyl or a fucosyl group. Fucosylated NFs were produced by an in vitro reaction with NodZ fucosyltransferase and purified. Their biological activity on pea was tested by measuring their capacity to stimulate the activity of a hydrolase that cleaves NFs. Non-modified and fucosylated NFs displayed this activity at nano- to picomolar concentrations, while a sulfated NF from Sinorhizobium meliloti was inactive. In an additional series of experiments, the stability of non-modified and fucosylated NFs in the presence of purified tobacco chitinases was compared. The presence of the fucosyl group affected the degradation rates and the accessibility of specific cleavage sites on the chitooligosaccharide backbone. These results suggest that the fucosyl group in NFs also weakens the interaction of NFs with certain chitinases or chitinase-related proteins in pea roots.

Effect of C6-Volatiles on Bioluminescent Plant Pathogens

HortScience ◽  
1998 ◽  
Vol 33 (3) ◽  
pp. 557d-557
Author(s):  
Jennifer Warr ◽  
Fenny Dane ◽  
Bob Ebel
Keyword(s):  
Biological Activity ◽  
Bacterial Growth ◽  
Xanthomonas Campestris ◽  
Plant Pathogens ◽  
Genetically Engineered ◽  
The Other ◽  
Computer Assisted ◽  
Signal Molecules ◽  
Low Concentrations

C6 volatile compounds are known to be produced by the plant upon pathogen attack or other stress-related events. The biological activity of many of these substances is poorly understood, but some might produce signal molecules important in host–pathogen interactions. In this research we explored the possibility that lipid-derived C6 volatiles have a direct effect on bacterial plant pathogens. To this purpose we used a unique tool, a bacterium genetically engineered to bioluminesce. Light-producing genes from a fish-associated bacterium were introduced into Xanthomonas campestris pv. campestris, enabling nondestructive detection of bacteria in vitro and in the plant with special computer-assisted camera equipment. The effects of different C6 volatiles (trans-2 hexanal, trans-2 hexen-1-ol and cis-3 hexenol) on growth of bioluminescent Xanthomonas campestris were investigated. Different volatile concentrations were used. Treatment with trans-2 hexanal appeared bactericidal at low concentrations (1% and 10%), while treatments with the other volatiles were not inhibitive to bacterial growth. The implications of these results with respect to practical use of trans-2 hexanal in pathogen susceptible and resistant plants will be discussed.

scholarly journals Rhizobial NodL O-Acetyl Transferase and NodS N-Methyl Transferase Functionally Interfere in Production of Modified Nod Factors

2001 ◽  
Vol 183 (11) ◽  
pp. 3408-3416 ◽  
Author(s):  
Isabel M. López-Lara ◽  
Dimitris Kafetzopoulos ◽  
Herman P. Spaink ◽  
Jane E. Thomas-Oates
Keyword(s):  
Nodule Formation ◽  
Signal Molecules ◽  
Nod Factors ◽  
Rhizobium Tropici ◽  
Methyl Transferase ◽  
Mesorhizobium Loti ◽  
Methyl Group ◽  
Rhizobial Species ◽  
Acetyl Group

ABSTRACT The products of the rhizobial nodulation genes are involved in the biosynthesis of lipochitin oligosaccharides (LCOs), which are host-specific signal molecules required for nodule formation. The presence of an O-acetyl group on C-6 of the nonreducingN-acetylglucosamine residue of LCOs is due to the enzymatic activity of NodL. Here we show that transfer of the nodLgene into four rhizobial species that all normally produce LCOs that are not modified on C-6 of the nonreducing terminal residue results in production of LCOs, the majority of which have an acetyl residue substituted on C-6. Surprisingly, in transconjugant strains ofMesorhizobium loti, Rhizobium etli, and Rhizobium tropici carrying nodL, such acetylation of LCOs prevents the endogenous nodS-dependent transfer of theN-methyl group that is found as a substituent of the acylated nitrogen atom. To study this interference betweennodL and nodS, we have cloned thenodS gene of M. loti and used its product in in vitro experiments in combination with purified NodL protein. It has previously been shown that a chitooligosaccharide N deacetylated on the nonreducing terminus (the so-called NodBC metabolite) is the preferred substrate for NodS as well as for NodL. Here we show that the NodBC metabolite, acetylated by NodL, is not used by the NodS protein as a substrate while the NodL protein can acetylate the NodBC metabolite that has been methylated by NodS.

scholarly journals A Sinorhizobium meliloti Lipopolysaccharide Mutant Altered in Cell Surface Sulfation

2002 ◽  
Vol 184 (23) ◽  
pp. 6681-6689 ◽  
Author(s):  
David H. Keating ◽  
Michael G. Willits ◽  
Sharon R. Long
Keyword(s):  
Cell Surface ◽  
Sinorhizobium Meliloti ◽  
Glucuronic Acid ◽  
Initial Study ◽  
Nodule Development ◽  
Nod Factor ◽  
Legume Symbiosis ◽  
Surface Polysaccharides

ABSTRACT The Rhizobium-legume symbiosis involves the formation of a novel plant organ, the nodule, in which intracellular bacteria reduce molecular dinitrogen in exchange for plant photosynthates. Nodule development requires a bacterial signal referred to as Nod factor, which in Sinorhizobium meliloti is a β-(1,4)-linked tetramer of N-acetylglucosamine containing N-acyl and O-acetyl modifications at the nonreducing end and a critical 6-O-sulfate at the reducing end. This sulfate modification requires the action of three gene products: nodH, which catalyzes the sulfonyl transfer, and nodPQ, which produce the activated form of sulfate, 3′-phosphoadenosine-5′-phosphosulfate. It was previously reported that S. meliloti cell surface polysaccharides are also covalently modified by sulfate in a reaction dependent on NodPQ. We have further characterized this unique form of bacterial carbohydrate modification. Our studies have determined that one of the nodPQ mutant strains used in the initial study of sulfation of cell surface harbored a second unlinked mutation. We cloned the gene affected by this mutation (referred to as lps-212) and found it to be an allele of lpsL, a gene previously predicted to encode a UDP-glucuronic acid epimerase. We demonstrated that lpsL encoded a UDP-glucuronic acid epimerase activity that was reduced in the lps-212 mutant. The lps-212 mutation resulted in an altered lipopolysaccharide structure that was reduced in sulfate modification in vitro and in vivo. Finally, we determined that the lps-212 mutation resulted in a reduced ability to elicit the formation of plant nodules and by altered infection thread structures that aborted prematurely.

scholarly journals Strain-Ecotype Specificity in Sinorhizobium meliloti-Medicago truncatula Symbiosis Is Correlated to Succinoglycan Oligosaccharide Structure

2007 ◽  
Vol 189 (21) ◽  
pp. 7733-7740 ◽  
Author(s):  
Senay Simsek ◽  
Tuula Ojanen-Reuhs ◽  
Samuel B. Stephens ◽  
Bradley L. Reuhs
Keyword(s):  
Medicago Truncatula ◽  
Sinorhizobium Meliloti ◽  
Donor Strain ◽  
Factor Activity ◽  
Nod Factors ◽  
Strain Specificity ◽  
Oligosaccharide Structure ◽  
Nod Factor ◽  
Structural Nature ◽  
Molecular Signals

ABSTRACT Molecular signals, including Nod factors and succinoglycan, are necessary for the establishment of nitrogen-fixing nodules (Fix+) in Medicago truncatula-Sinorhizobium meliloti symbiosis. This report shows that M. truncatula-S. meliloti interactions involve ecotype-strain specificity, as S. meliloti Rm41 and NRG247 are Fix+ (compatible) on M. truncatula A20 and Fix− (incompatible) on M. truncatula A17, the Fix phenotypes are reversed with S. meliloti NRG185 and NRG34, and there is a correlation between the host specificity and succinoglycan oligosaccharide structure. S. meliloti NRG185 produces oligosaccharides that are almost fully succinylated, with two succinate groups per subunit, whereas the oligosaccharides produced by S. meliloti Rm41 include many nonsuccinylated subunits, as well as subunits with a single succinate group and others with malate. The results of this study demonstrated the following: (i) incompatibility is not a consequence of an avirulence factor or lack of Nod factor activity; (ii) the Fix+ phenotypes are succinoglycan dependent; (iii) there is structural variability in the succinoglycan oligosaccharide populations between S. meliloti strains; (iv) the structural nature of the succinoglycan oligosaccharides is correlated to compatibility; most importantly, (v) an S. meliloti Rm41 derivative, carrying exo genes from an M. truncatula A17-compatible strain, produced a modified population of succinoglycan oligosaccharides (similar to the donor strain) and was Fix+ on A17.

scholarly journals l-Canavanine Made by Medicago sativa Interferes with Quorum Sensing in Sinorhizobium meliloti

2005 ◽  
Vol 187 (24) ◽  
pp. 8427-8436 ◽  
Author(s):  
Neela D. Keshavan ◽  
Puneet K. Chowdhary ◽  
Donovan C. Haines ◽  
Juan E. González
Keyword(s):  
Quorum Sensing ◽  
Medicago Sativa ◽  
Sinorhizobium Meliloti ◽  
Signal Molecules ◽  
Bacterial Strains ◽  
Signal Molecule ◽  
Gram Negative ◽  
Bacterial Quorum Sensing ◽  
Bacterial Quorum ◽  
Regulated Gene Expression

ABSTRACT Sinorhizobium meliloti is a gram-negative soil bacterium, capable of establishing a nitrogen-fixing symbiosis with its legume host, alfalfa (Medicago sativa). Quorum sensing plays a crucial role in this symbiosis, where it influences the nodulation process and the synthesis of the symbiotically important exopolysaccharide II (EPS II). S. meliloti has three quorum-sensing systems (Sin, Tra, and Mel) that use N-acyl homoserine lactones as their quorum-sensing signal molecule. Increasing evidence indicates that certain eukaryotic hosts involved in symbiotic or pathogenic relationships with gram-negative bacteria produce quorum-sensing-interfering (QSI) compounds that can cross-communicate with the bacterial quorum-sensing system. Our studies of alfalfa seed exudates suggested the presence of multiple signal molecules capable of interfering with quorum-sensing-regulated gene expression in different bacterial strains. In this work, we choose one of these QSI molecules (SWI) for further characterization. SWI inhibited violacein production, a phenotype that is regulated by quorum sensing in Chromobacterium violaceum. In addition, this signal molecule also inhibits the expression of the S. meliloti exp genes, responsible for the production of EPS II, a quorum-sensing-regulated phenotype. We identified this molecule as l-canavanine, an arginine analog, produced in large quantities by alfalfa and other legumes.

scholarly journals Rapid Colorimetric Quantification of Lipo-chitooligosaccharides from Mesorhizobium loti and Sinorhizobium meliloti

2002 ◽  
Vol 15 (9) ◽  
pp. 859-865
Author(s):  
Joachim Goedhart ◽  
Jean-Jacques Bono ◽  
Theodorus W. J. Gadella
Keyword(s):  
Methylene Blue ◽  
Organic Phase ◽  
Sinorhizobium Meliloti ◽  
Sodium Dodecyl ◽  
Ion Pair ◽  
Nod Factors ◽  
Two Phase ◽  
Nod Factor ◽  
Mesorhizobium Loti ◽  
Hydrolysis Of

Nod factors are lipids with a chitinlike headgroup produced by gram-negative Rhizobium bacteria. These lipo-chitooligosaccharides (LCOs) are essential signaling molecules for accomplishing symbiosis between the bacteria and roots of legume plants. Despite their important role in the Rhizobium-legume interaction, no fast and sensitive Nod factor quantification methods exist. Here, we report two different quantification methods. The first is based on the enzymatic hydrolysis of Nod factors to release N-acetylglucosamine (GlcNAc), which can subsequently be quantified. It is shown that the degrading enzyme, glusulase, releases exactly two GlcNAc units per pentameric nodulation factor from Mesorhizobium loti factor, allowing quantification of LCOs from Mesorhizobium loti. The second method is based on a specific type of Nod factors that are sulfated on the reducing GlcNAc, allowing quantification analogous to the quantification of sulfolipids. Here, a two-phase extraction method is used in the presence of methylene blue, which specifically forms an ion pair with sulfated lipids. The blue ion pair partitions into the organic phase, after which the methylene blue signal can be quantified. To enable Nod factor quantification with this method, the organic phase was modified and the partitioning was evaluated using fluorescent and radiolabeled sulfated Nod factors. It is shown that sulfated LCOs can be quantified with this method, using sodium dodecyl sulfate for calibration. Both methods allow Nod factor quantification in parallel enabling a fast and easy detection of nanomole quantities of Nod factors. Accurate Nod factor quantification will be crucial for characterization and cross-comparison of the affinity for Nod factors of newly identified Nod factor binding proteins or putative Nod factor receptors.

scholarly journals Synthesis of New Heterocyclic Compounds Derived from Anthrone and Evaluation of Their Biological Activity

2008 ◽  
Vol 5 (4) ◽  
pp. 627-637
Author(s):  
Baghdad Science Journal
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Biological Activity ◽  
Heterocyclic Compounds ◽  
Heterocyclic Amines ◽  
Pathogenic Microorganism ◽  
Standard Strain ◽  
Chemical Structures ◽  
Heterocyclic Derivatives

In this research, Schiff bases derived from the reaction of anthrone with different heterocyclic amines have been described. The resulted Schiff base compounds were reacted with various nucleophiles in order to obtain new heterocyclic derivatives. Chemical structures of all products were confirmed by IR, 1H-, 13C-NMR spectral data and elemental analysis. All synthesized compounds were in vitro tested against a standard strain of pathogenic microorganism including Gram +ve bacteria (Staphylococcus aureus), Gram –ve bacteria (Escherichia coli), and fungi (Candida albicans).

scholarly journals Sinorhizobium meliloti Sulfotransferase That Modifies Lipopolysaccharide

2004 ◽  
Vol 186 (13) ◽  
pp. 4168-4176 ◽  
Author(s):  
Glen E. Cronan ◽  
David H. Keating
Keyword(s):  
Cell Surface ◽  
Sinorhizobium Meliloti ◽  
Capsular Polysaccharide ◽  
Genomic Sequence ◽  
Wild Type ◽  
Plant Structure ◽  
Plant Host ◽  
Nod Factor

ABSTRACT Sinorhizobium meliloti is a gram-negative soil bacterium found either in free-living form or as a nitrogen-fixing endosymbiont of a plant structure called the nodule. Symbiosis between S. meliloti and its plant host alfalfa is dependent on bacterial transcription of nod genes, which encode the enzymes responsible for synthesis of Nod factor. S. meliloti Nod factor is a lipochitooligosaccharide that undergoes a sulfate modification essential for its biological activity. Sulfate also modifies the carbohydrate substituents of the bacterial cell surface, including lipopolysaccharide (LPS) and capsular polysaccharide (K-antigen) (R. A. Cedergren, J. Lee, K. L. Ross, and R. I. Hollingsworth, Biochemistry 34:4467-4477, 1995). We utilized the genomic sequence of S. meliloti to identify an open reading frame, SMc04267 (which we now propose to name lpsS), which encodes an LPS sulfotransferase activity. We expressed LpsS in Escherichia coli and demonstrated that the purified protein functions as an LPS sulfotransferase. Mutants lacking LpsS displayed an 89% reduction in LPS sulfotransferase activity in vitro. However, lpsS mutants retain approximately wild-type levels of sulfated LPS when assayed in vivo, indicating the presence of an additional LPS sulfotransferase activity(ies) in S. meliloti that can compensate for the loss of LpsS. The lpsS mutant did show reduced LPS sulfation, compared to that of the wild type, under conditions that promote nod gene expression, and it elicited a greater number of nodules than did the wild type during symbiosis with alfalfa. These results suggest that sulfation of cell surface polysaccharides and Nod factor may compete for a limiting pool of intracellular sulfate and that LpsS is required for optimal LPS sulfation under these conditions.

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