Jolanta Kutkowska | Maria Curie - Skłodowska University (original) (raw)

Papers by Jolanta Kutkowska

Journal of Molecular Structure, 2012

BMC Microbiology, 2010

Background: Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that el... more Background: Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodules on roots of host plants Trifolium spp. Bacterial surface polysaccharides are crucial for establishment of a successful symbiosis with legumes that form indeterminate-type nodules, such as Trifolium, Pisum, Vicia, and Medicago spp. and aid the bacterium in withstanding osmotic and other environmental stresses. Recently, the R. leguminosarum bv. trifolii RosR regulatory protein which controls exopolysaccharide production has been identified and characterized. Results: In this work, we extend our earlier studies to the characterization of rosR mutants which exhibit pleiotropic phenotypes. The mutants produce three times less exopolysaccharide than the wild type, and the lowmolecular-weight fraction in that polymer is greatly reduced. Mutation in rosR also results in quantitative alterations in the polysaccharide constituent of lipopolysaccharide. The rosR mutants are more sensitive to surface-active detergents, antibiotics of the beta-lactam group and some osmolytes, indicating changes in the bacterial membranes. In addition, the rosR mutants exhibit significant decrease in motility and form a biofilm on plastic surfaces, which differs significantly in depth, architecture, and bacterial viability from that of the wild type. The most striking effect of rosR mutation is the considerably decreased attachment and colonization of root hairs, indicating that the mutation affects the first stage of the invasion process. Infection threads initiate at a drastically reduced rate and frequently abort before they reach the base of root hairs. Although these mutants form nodules on clover, they are unable to fix nitrogen and are outcompeted by the wild type in mixed inoculations, demonstrating that functional rosR is important for competitive nodulation. Conclusions: This report demonstrates the significant role RosR regulatory protein plays in bacterial stress adaptation and in the symbiotic relationship between clover and R. leguminosarum bv. trifolii 24.2.

Acta poloniae pharmaceutica

In this study, some O-benzyl (benzofuran-2-yl)ethan-1-one ether oximes were synthesized starting ... more In this study, some O-benzyl (benzofuran-2-yl)ethan-1-one ether oximes were synthesized starting from 2-acetylbenzofuran. The structure elucidation of the compounds was performed by IR, 1H-NMR and 13C-NMR spectra. Antimicrobial activities of the compounds were examined and notable activity was observed.

Tetrahedron Letters, 2010

An analysis of the products of the reaction of N 3 -phenylbenzamidrazone with cis-1,2-cyclohexane... more An analysis of the products of the reaction of N 3 -phenylbenzamidrazone with cis-1,2-cyclohexanedicarboxylic anhydride at various temperatures is presented. The identification of the reaction products is carried on with the support of computational techniques. The most stable conformers of the isoindole and triazole derivatives are found within the DFT approach. The theoretical calculations reveal the possible structure of a triazole derivative not available experimentally because of the presence of two diastereoisomers of equal energy.

Systematic and Applied Microbiology, 2000

Lipopolysaccharides (LPSs) isolated from seven strains of Mesorhizobium were studied for the pres... more Lipopolysaccharides (LPSs) isolated from seven strains of Mesorhizobium were studied for the presence of fatty acids with particular attention for 27-oxooctacosanoic acid and 4-oxo fatty acids. The LPSs from all analysed strains contained various amounts of 27-oxo-28:0 and all of them, with the exception of Mesorhizobium tianshanense, contained also 4-oxo fatty acids (4-oxo-20:0, 4-oxo-i-21:0, 4-oxo-22:0). The group of amide-linked fatty acids consisted of a wide range of 3-hydroxylated and 4-oxo fatty acids whereas all the nonpolar as well as the (m-l) hydroxylated long-chain acids and the 27-oxo-28:0 fatty acids were ester-linked. The characteristic spectrum of 3-hydroxy fatty acids and presence of 27-0H-28:0 as well as 27-oxo-28:0 acid in LPSs of Mesorhizobium showed that these strains were closely related. Therefore the lipid A fatty acid pattern could be a useful chemotaxonomic marker which helps to isolate the Mesorhizobium group from rhizobium bacteria during the classification process.

Journal of Molecular Structure, 2012

Biochemistry (Moscow), 2011

Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant... more Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant Rt120 in the pssBpssA intergenic region as well as degraded polysaccharides (DPS) derived from the LPS were elucidated in terms of their chemical composition and biological activities. The polysaccharide portions were examined by methylation analysis, MALDI-TOF mass spectrometry, and (1)H NMR spectroscopy. A high molecular mass carbohydrate fraction obtained from Rt120 DPS by Sephadex G-50 gel chromatography was composed mainly of L-rhamnose, 6-deoxy-L-talose, D-galactose, and D-galacturonic acid, whereas that from RtTA1 DPS contained L-fucose, 2-acetamido-2,6-dideoxy-D-glucose, D-galacturonic acid, 3-deoxy-3-methylaminofucose, D-glucose, D-glucuronic acid, and heptose. Relative intensities of the major (1)H NMR signals for O-acetyl and N-acetyl groups were 1 : 0.8 and 1 : 1.24 in DPS of Rt120 and RtTA1, respectively. The intact mutant LPS exhibited a twice higher lethal toxicity than the wild type LPS. A higher in vivo production of TNFα and IL-6 after induction of mice with Rt120 LPS correlated with the toxicity, although the mutant LPS induced the secretion of IL-1β and IFNγ more weakly than RtTA1 LPS. A polysaccharide obtained by gel chromatography on Bio-Gel P-4 of the high molecular mass material from Rt120 had a toxic effect on tumor HeLa cells but was inactive against the normal human skin fibroblast cell line. The polysaccharide from RtTA1 was inactive against either cell line. The potent inhibitory effect of the mutant DPS on tumor HeLa cells seems to be related with the differences in sugar composition.

Archives of Microbiology, Sep 3, 2004

Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the ... more Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the pssD, pssP, pssT and pssO genes and altered in exopolysaccharide (EPS) synthesis were investigated. EPS-deficient mutants were also changed in lipopolysaccharide structure. All mutants exhibited varied sensitivities to detergents, ethanol and antibiotics, thus indicating changes in bacterial membrane integrity. Using pss mutants marked with the gusA gene, EPS-deficient mutants were found to have abnormalities in nodule development and to provoke severe plant defence reactions. The pss mutants that produced altered quantities of EPS with a changed degree of polymerisation generally occupied the younger developmental zones of the nodules and elicited moderate plant defence reactions.

PLoS ONE, 2014

Production of extracellular polysaccharides is a complex process engaging proteins localized in d... more Production of extracellular polysaccharides is a complex process engaging proteins localized in different subcellular compartments, yet communicating with each other or even directly interacting in multicomponent complexes. Proteins involved in polymerization and transport of exopolysaccharide (EPS) in Rhizobium leguminosarum are encoded within the chromosomal Pss-I cluster. However, genes implicated in polysaccharide synthesis are common in rhizobia, with several homologues of pss genes identified in other regions of the R. leguminosarum genome. One such region is chromosomally located Pss-II encoding proteins homologous to known components of the Wzx/Wzy-dependent polysaccharide synthesis and transport systems. The pssP2 gene encodes a protein similar to polysaccharide co-polymerases involved in determination of the length of polysaccharide chains in capsule and O-antigen biosynthesis. In this work, a mutant with a disrupted pssP2 gene was constructed and its capabilities to produce EPS and enter into a symbiotic relationship with clover were studied. The pssP2 mutant, while not altered in lipopolysaccharide (LPS), displayed changes in molecular mass distribution profile of EPS. Lack of the full-length PssP2 protein resulted in a reduction of high molecular weight EPS, yet polymerized to a longer length than in the RtTA1 wild type. The mutant strain was also more efficient in symbiotic performance. The functional interrelation between PssP2 and proteins encoded within the Pss-I region was further supported by data from bacterial two-hybrid assays providing evidence for PssP2 interactions with PssT polymerase, as well as glycosyltransferase PssC. A possible role for PssP2 in a complex involved in EPS chain-length determination is discussed.

Microbiological Research, 2009

Mesorhizobium loti NZP2213.1 mutant obtained after random Tn5 mutagenesis of M. loti NZP2213 was ... more Mesorhizobium loti NZP2213.1 mutant obtained after random Tn5 mutagenesis of M. loti NZP2213 was inefficient in nitrogen fixation on Lotus corniculatus. The transposon insertion was located within an ORF with a sequence similarity to a putative glycosyl transferase from Caulobacter crescentus. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the mutant produced LPS of the same O-chain length but only half of the entire smooth LPS, compared to that of the parental strain. A greater diversity of the anomeric region as determined by NMR spectroscopy, reflected structural differences in the mutant repeating units represented by 6-deoxytalose, 2-OAc-6-deoxytalose, and 2-OMe-6deoxytalose. In contrast to the completely O-acetylated 6-deoxytalose in wild-type OPS only partial O-acetylation was found in the mutant. The decrease of the LPS species with O-chains seems to be correlated with 6-deoxytalose deficiency. Microscopic examination of the nodules induced by the mutant revealed disturbances in infection thread development and premature senescence of symbiosomes. The impairment of mutant-induced symbiosomes to sustain latter stages of symbiosis could be a consequence of the decreased ratio of the hydrophobic to the hydrophilic LPSs. J 5,6 3 J H,H (Hz) a 2, d 3.5, dd 3.5, t $1, dd 6.5, m a d, dublet; dd, double dublet; t, triplet; m, multiplet.

Journal of Plant Physiology, 2001

A Rhizobium leguminosarum bv. trifolii Tn5 transposon mutant deficient in exopolysaccharide biosy... more A Rhizobium leguminosarum bv. trifolii Tn5 transposon mutant deficient in exopolysaccharide biosynthesis was found to form non-nitrogen fixing nodules on clover. Root nodules induced by the mutant contained aberrant infection threads and few bacteroids. Sequence analysis of the transposon insertion site localized the mutation in the pssB-pssA intergenic region affecting the exopolysaccharide biosynthesis. The mutant also showed decreased sensitivity to SDS and deoxycholate and displayed a changed lipopolysaccharide (LPS) banding pattern compared to the wild-type strain TA1. The alteration in the O-polysaccharide part of LPS was confirmed by Western immunoblotting with polyclonal antibodies. LPS preparations of strain TA1 and the mutant strain only reacted with their homologous sera. The common epitopes in LPS from bacteroids and free-living rhizobia were revealed by immunogold assay. The results of this study indicate that the pssB-pssA region of R. leguminosarum bv. trifolii is important for the polysaccharide synthesis.

Synthesis, crystal structure and biological activities of a novel amidrazone derivative and its copper(II) complex — A potential antitumor drug

Journal of Inorganic Biochemistry, 2012

Immunobiology, 2000

The lethality and eytokines-inducing activity of lipopolysaccharides (LPS) obtained from nodulati... more The lethality and eytokines-inducing activity of lipopolysaccharides (LPS) obtained from nodulating bacteria, Rhizobium leguminosarum and Mesorhizobium loti, were compared to those of Salmonella typhimurium LPS. The activity of R. leguminosarum LPS was almost comparable to Salmonella endotoxin in terms of lethality, Limulus lysate gelating activity and in vivo tumor necrosis factor-a (TNF-a), interleukin-l~(IL-l~), interleukin-6 (IL-6) and interferon-y (IFN-y) induction capacity. In contrast to high lethal toxicity of Rhizobium LPS, the lethality of LPS isolated from Mesorhizobium loti was more than 10 3 -fold lower. Weak lethality of LPS from Mesorhizobium correlated with low capacity of this LPS to induce TNF-a, IL-1~, IL-6 and IFN-y both in vivo and in vitro in murine splenoeytes.

Carbohydrate Research, 2009

The O-polysaccharide of Mesorhizobium loti HAMBI 1148 was obtained by mild acid degradation of th... more The O-polysaccharide of Mesorhizobium loti HAMBI 1148 was obtained by mild acid degradation of the lipopolysaccharide and studied by sugar and methylation analyses, Smith degradation, and 1 H and 13 C NMR spectroscopies, including 2D 1 H/ 1 H COSY, TOCSY, ROESY, and H-detected 1 H/ 13 C HSQC experiments. The O-polysaccharide was found to have a branched hexasaccharide-repeating unit of the following structure:

Biological activity of (Lipo)polysaccharides of the exopolysaccharide-deficient mutant Rt120 derived from Rhizobium leguminosarum bv. trifolii strain TA1

Biochemistry (Moscow), 2011

Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant... more Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant Rt120 in the pssBpssA intergenic region as well as degraded polysaccharides (DPS) derived from the LPS were elucidated in terms of their chemical composition and biological activities. The polysaccharide portions were examined by methylation analysis, MALDI-TOF mass spectrometry, and (1)H NMR spectroscopy. A high molecular mass carbohydrate fraction obtained from Rt120 DPS by Sephadex G-50 gel chromatography was composed mainly of L-rhamnose, 6-deoxy-L-talose, D-galactose, and D-galacturonic acid, whereas that from RtTA1 DPS contained L-fucose, 2-acetamido-2,6-dideoxy-D-glucose, D-galacturonic acid, 3-deoxy-3-methylaminofucose, D-glucose, D-glucuronic acid, and heptose. Relative intensities of the major (1)H NMR signals for O-acetyl and N-acetyl groups were 1 : 0.8 and 1 : 1.24 in DPS of Rt120 and RtTA1, respectively. The intact mutant LPS exhibited a twice higher lethal toxicity than the wild type LPS. A higher in vivo production of TNFα and IL-6 after induction of mice with Rt120 LPS correlated with the toxicity, although the mutant LPS induced the secretion of IL-1β and IFNγ more weakly than RtTA1 LPS. A polysaccharide obtained by gel chromatography on Bio-Gel P-4 of the high molecular mass material from Rt120 had a toxic effect on tumor HeLa cells but was inactive against the normal human skin fibroblast cell line. The polysaccharide from RtTA1 was inactive against either cell line. The potent inhibitory effect of the mutant DPS on tumor HeLa cells seems to be related with the differences in sugar composition.

Archives of Microbiology, 2004

Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the ... more Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the pssD, pssP, pssT and pssO genes and altered in exopolysaccharide (EPS) synthesis were investigated. EPS-deficient mutants were also changed in lipopolysaccharide structure. All mutants exhibited varied sensitivities to detergents, ethanol and antibiotics, thus indicating changes in bacterial membrane integrity. Using pss mutants marked with the gusA gene, EPS-deficient mutants were found to have abnormalities in nodule development and to provoke severe plant defence reactions. The pss mutants that produced altered quantities of EPS with a changed degree of polymerisation generally occupied the younger developmental zones of the nodules and elicited moderate plant defence reactions.

Archiv der Pharmazie, 2012

A series of novel compounds were synthesized in reactions of N 3 -substituted amidrazones with ci... more A series of novel compounds were synthesized in reactions of N 3 -substituted amidrazones with cis-1,2cyclohexanedicarboxylic anhydride: linear, isoindole, and triazole derivatives. All new structures were confirmed by H 1 NMR and IR spectrometry as well as elemental analysis. Potential biological effects of new compounds were predicted with the Prediction of Activity Spectra for Substances (PASS) program. Antiviral, antibacterial, analgesic, and anti-inflammatory activities were experimentally verified.

BMC Microbiology

Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodule... more Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodules on roots of host plants Trifolium spp. Bacterial surface polysaccharides are crucial for establishment of a successful symbiosis with legumes that form indeterminate-type nodules, such as Trifolium, Pisum, Vicia, and Medicago spp. and aid the bacterium in withstanding osmotic and other environmental stresses. Recently, the R. leguminosarum bv. trifolii RosR regulatory protein which controls exopolysaccharide production has been identified and characterized. In this work, we extend our earlier studies to the characterization of rosR mutants which exhibit pleiotropic phenotypes. The mutants produce three times less exopolysaccharide than the wild type, and the low-molecular-weight fraction in that polymer is greatly reduced. Mutation in rosR also results in quantitative alterations in the polysaccharide constituent of lipopolysaccharide. The rosR mutants are more sensitive to surface-act...

Journal of Molecular Structure, 2012

BMC Microbiology, 2010

Background: Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that el... more Background: Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodules on roots of host plants Trifolium spp. Bacterial surface polysaccharides are crucial for establishment of a successful symbiosis with legumes that form indeterminate-type nodules, such as Trifolium, Pisum, Vicia, and Medicago spp. and aid the bacterium in withstanding osmotic and other environmental stresses. Recently, the R. leguminosarum bv. trifolii RosR regulatory protein which controls exopolysaccharide production has been identified and characterized. Results: In this work, we extend our earlier studies to the characterization of rosR mutants which exhibit pleiotropic phenotypes. The mutants produce three times less exopolysaccharide than the wild type, and the lowmolecular-weight fraction in that polymer is greatly reduced. Mutation in rosR also results in quantitative alterations in the polysaccharide constituent of lipopolysaccharide. The rosR mutants are more sensitive to surface-active detergents, antibiotics of the beta-lactam group and some osmolytes, indicating changes in the bacterial membranes. In addition, the rosR mutants exhibit significant decrease in motility and form a biofilm on plastic surfaces, which differs significantly in depth, architecture, and bacterial viability from that of the wild type. The most striking effect of rosR mutation is the considerably decreased attachment and colonization of root hairs, indicating that the mutation affects the first stage of the invasion process. Infection threads initiate at a drastically reduced rate and frequently abort before they reach the base of root hairs. Although these mutants form nodules on clover, they are unable to fix nitrogen and are outcompeted by the wild type in mixed inoculations, demonstrating that functional rosR is important for competitive nodulation. Conclusions: This report demonstrates the significant role RosR regulatory protein plays in bacterial stress adaptation and in the symbiotic relationship between clover and R. leguminosarum bv. trifolii 24.2.

Acta poloniae pharmaceutica

In this study, some O-benzyl (benzofuran-2-yl)ethan-1-one ether oximes were synthesized starting ... more In this study, some O-benzyl (benzofuran-2-yl)ethan-1-one ether oximes were synthesized starting from 2-acetylbenzofuran. The structure elucidation of the compounds was performed by IR, 1H-NMR and 13C-NMR spectra. Antimicrobial activities of the compounds were examined and notable activity was observed.

Tetrahedron Letters, 2010

An analysis of the products of the reaction of N 3 -phenylbenzamidrazone with cis-1,2-cyclohexane... more An analysis of the products of the reaction of N 3 -phenylbenzamidrazone with cis-1,2-cyclohexanedicarboxylic anhydride at various temperatures is presented. The identification of the reaction products is carried on with the support of computational techniques. The most stable conformers of the isoindole and triazole derivatives are found within the DFT approach. The theoretical calculations reveal the possible structure of a triazole derivative not available experimentally because of the presence of two diastereoisomers of equal energy.

Systematic and Applied Microbiology, 2000

Lipopolysaccharides (LPSs) isolated from seven strains of Mesorhizobium were studied for the pres... more Lipopolysaccharides (LPSs) isolated from seven strains of Mesorhizobium were studied for the presence of fatty acids with particular attention for 27-oxooctacosanoic acid and 4-oxo fatty acids. The LPSs from all analysed strains contained various amounts of 27-oxo-28:0 and all of them, with the exception of Mesorhizobium tianshanense, contained also 4-oxo fatty acids (4-oxo-20:0, 4-oxo-i-21:0, 4-oxo-22:0). The group of amide-linked fatty acids consisted of a wide range of 3-hydroxylated and 4-oxo fatty acids whereas all the nonpolar as well as the (m-l) hydroxylated long-chain acids and the 27-oxo-28:0 fatty acids were ester-linked. The characteristic spectrum of 3-hydroxy fatty acids and presence of 27-0H-28:0 as well as 27-oxo-28:0 acid in LPSs of Mesorhizobium showed that these strains were closely related. Therefore the lipid A fatty acid pattern could be a useful chemotaxonomic marker which helps to isolate the Mesorhizobium group from rhizobium bacteria during the classification process.

Journal of Molecular Structure, 2012

Biochemistry (Moscow), 2011

Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant... more Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant Rt120 in the pssBpssA intergenic region as well as degraded polysaccharides (DPS) derived from the LPS were elucidated in terms of their chemical composition and biological activities. The polysaccharide portions were examined by methylation analysis, MALDI-TOF mass spectrometry, and (1)H NMR spectroscopy. A high molecular mass carbohydrate fraction obtained from Rt120 DPS by Sephadex G-50 gel chromatography was composed mainly of L-rhamnose, 6-deoxy-L-talose, D-galactose, and D-galacturonic acid, whereas that from RtTA1 DPS contained L-fucose, 2-acetamido-2,6-dideoxy-D-glucose, D-galacturonic acid, 3-deoxy-3-methylaminofucose, D-glucose, D-glucuronic acid, and heptose. Relative intensities of the major (1)H NMR signals for O-acetyl and N-acetyl groups were 1 : 0.8 and 1 : 1.24 in DPS of Rt120 and RtTA1, respectively. The intact mutant LPS exhibited a twice higher lethal toxicity than the wild type LPS. A higher in vivo production of TNFα and IL-6 after induction of mice with Rt120 LPS correlated with the toxicity, although the mutant LPS induced the secretion of IL-1β and IFNγ more weakly than RtTA1 LPS. A polysaccharide obtained by gel chromatography on Bio-Gel P-4 of the high molecular mass material from Rt120 had a toxic effect on tumor HeLa cells but was inactive against the normal human skin fibroblast cell line. The polysaccharide from RtTA1 was inactive against either cell line. The potent inhibitory effect of the mutant DPS on tumor HeLa cells seems to be related with the differences in sugar composition.

Archives of Microbiology, Sep 3, 2004

Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the ... more Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the pssD, pssP, pssT and pssO genes and altered in exopolysaccharide (EPS) synthesis were investigated. EPS-deficient mutants were also changed in lipopolysaccharide structure. All mutants exhibited varied sensitivities to detergents, ethanol and antibiotics, thus indicating changes in bacterial membrane integrity. Using pss mutants marked with the gusA gene, EPS-deficient mutants were found to have abnormalities in nodule development and to provoke severe plant defence reactions. The pss mutants that produced altered quantities of EPS with a changed degree of polymerisation generally occupied the younger developmental zones of the nodules and elicited moderate plant defence reactions.

PLoS ONE, 2014

Production of extracellular polysaccharides is a complex process engaging proteins localized in d... more Production of extracellular polysaccharides is a complex process engaging proteins localized in different subcellular compartments, yet communicating with each other or even directly interacting in multicomponent complexes. Proteins involved in polymerization and transport of exopolysaccharide (EPS) in Rhizobium leguminosarum are encoded within the chromosomal Pss-I cluster. However, genes implicated in polysaccharide synthesis are common in rhizobia, with several homologues of pss genes identified in other regions of the R. leguminosarum genome. One such region is chromosomally located Pss-II encoding proteins homologous to known components of the Wzx/Wzy-dependent polysaccharide synthesis and transport systems. The pssP2 gene encodes a protein similar to polysaccharide co-polymerases involved in determination of the length of polysaccharide chains in capsule and O-antigen biosynthesis. In this work, a mutant with a disrupted pssP2 gene was constructed and its capabilities to produce EPS and enter into a symbiotic relationship with clover were studied. The pssP2 mutant, while not altered in lipopolysaccharide (LPS), displayed changes in molecular mass distribution profile of EPS. Lack of the full-length PssP2 protein resulted in a reduction of high molecular weight EPS, yet polymerized to a longer length than in the RtTA1 wild type. The mutant strain was also more efficient in symbiotic performance. The functional interrelation between PssP2 and proteins encoded within the Pss-I region was further supported by data from bacterial two-hybrid assays providing evidence for PssP2 interactions with PssT polymerase, as well as glycosyltransferase PssC. A possible role for PssP2 in a complex involved in EPS chain-length determination is discussed.

Microbiological Research, 2009

Mesorhizobium loti NZP2213.1 mutant obtained after random Tn5 mutagenesis of M. loti NZP2213 was ... more Mesorhizobium loti NZP2213.1 mutant obtained after random Tn5 mutagenesis of M. loti NZP2213 was inefficient in nitrogen fixation on Lotus corniculatus. The transposon insertion was located within an ORF with a sequence similarity to a putative glycosyl transferase from Caulobacter crescentus. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the mutant produced LPS of the same O-chain length but only half of the entire smooth LPS, compared to that of the parental strain. A greater diversity of the anomeric region as determined by NMR spectroscopy, reflected structural differences in the mutant repeating units represented by 6-deoxytalose, 2-OAc-6-deoxytalose, and 2-OMe-6deoxytalose. In contrast to the completely O-acetylated 6-deoxytalose in wild-type OPS only partial O-acetylation was found in the mutant. The decrease of the LPS species with O-chains seems to be correlated with 6-deoxytalose deficiency. Microscopic examination of the nodules induced by the mutant revealed disturbances in infection thread development and premature senescence of symbiosomes. The impairment of mutant-induced symbiosomes to sustain latter stages of symbiosis could be a consequence of the decreased ratio of the hydrophobic to the hydrophilic LPSs. J 5,6 3 J H,H (Hz) a 2, d 3.5, dd 3.5, t $1, dd 6.5, m a d, dublet; dd, double dublet; t, triplet; m, multiplet.

Journal of Plant Physiology, 2001

A Rhizobium leguminosarum bv. trifolii Tn5 transposon mutant deficient in exopolysaccharide biosy... more A Rhizobium leguminosarum bv. trifolii Tn5 transposon mutant deficient in exopolysaccharide biosynthesis was found to form non-nitrogen fixing nodules on clover. Root nodules induced by the mutant contained aberrant infection threads and few bacteroids. Sequence analysis of the transposon insertion site localized the mutation in the pssB-pssA intergenic region affecting the exopolysaccharide biosynthesis. The mutant also showed decreased sensitivity to SDS and deoxycholate and displayed a changed lipopolysaccharide (LPS) banding pattern compared to the wild-type strain TA1. The alteration in the O-polysaccharide part of LPS was confirmed by Western immunoblotting with polyclonal antibodies. LPS preparations of strain TA1 and the mutant strain only reacted with their homologous sera. The common epitopes in LPS from bacteroids and free-living rhizobia were revealed by immunogold assay. The results of this study indicate that the pssB-pssA region of R. leguminosarum bv. trifolii is important for the polysaccharide synthesis.

Synthesis, crystal structure and biological activities of a novel amidrazone derivative and its copper(II) complex — A potential antitumor drug

Journal of Inorganic Biochemistry, 2012

Immunobiology, 2000

The lethality and eytokines-inducing activity of lipopolysaccharides (LPS) obtained from nodulati... more The lethality and eytokines-inducing activity of lipopolysaccharides (LPS) obtained from nodulating bacteria, Rhizobium leguminosarum and Mesorhizobium loti, were compared to those of Salmonella typhimurium LPS. The activity of R. leguminosarum LPS was almost comparable to Salmonella endotoxin in terms of lethality, Limulus lysate gelating activity and in vivo tumor necrosis factor-a (TNF-a), interleukin-l~(IL-l~), interleukin-6 (IL-6) and interferon-y (IFN-y) induction capacity. In contrast to high lethal toxicity of Rhizobium LPS, the lethality of LPS isolated from Mesorhizobium loti was more than 10 3 -fold lower. Weak lethality of LPS from Mesorhizobium correlated with low capacity of this LPS to induce TNF-a, IL-1~, IL-6 and IFN-y both in vivo and in vitro in murine splenoeytes.

Carbohydrate Research, 2009

The O-polysaccharide of Mesorhizobium loti HAMBI 1148 was obtained by mild acid degradation of th... more The O-polysaccharide of Mesorhizobium loti HAMBI 1148 was obtained by mild acid degradation of the lipopolysaccharide and studied by sugar and methylation analyses, Smith degradation, and 1 H and 13 C NMR spectroscopies, including 2D 1 H/ 1 H COSY, TOCSY, ROESY, and H-detected 1 H/ 13 C HSQC experiments. The O-polysaccharide was found to have a branched hexasaccharide-repeating unit of the following structure:

Biological activity of (Lipo)polysaccharides of the exopolysaccharide-deficient mutant Rt120 derived from Rhizobium leguminosarum bv. trifolii strain TA1

Biochemistry (Moscow), 2011

Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant... more Lipopolysaccharides (LPS) from Rhizobium leguminosarum biovar trifolii TA1 (RtTA1) and its mutant Rt120 in the pssBpssA intergenic region as well as degraded polysaccharides (DPS) derived from the LPS were elucidated in terms of their chemical composition and biological activities. The polysaccharide portions were examined by methylation analysis, MALDI-TOF mass spectrometry, and (1)H NMR spectroscopy. A high molecular mass carbohydrate fraction obtained from Rt120 DPS by Sephadex G-50 gel chromatography was composed mainly of L-rhamnose, 6-deoxy-L-talose, D-galactose, and D-galacturonic acid, whereas that from RtTA1 DPS contained L-fucose, 2-acetamido-2,6-dideoxy-D-glucose, D-galacturonic acid, 3-deoxy-3-methylaminofucose, D-glucose, D-glucuronic acid, and heptose. Relative intensities of the major (1)H NMR signals for O-acetyl and N-acetyl groups were 1 : 0.8 and 1 : 1.24 in DPS of Rt120 and RtTA1, respectively. The intact mutant LPS exhibited a twice higher lethal toxicity than the wild type LPS. A higher in vivo production of TNFα and IL-6 after induction of mice with Rt120 LPS correlated with the toxicity, although the mutant LPS induced the secretion of IL-1β and IFNγ more weakly than RtTA1 LPS. A polysaccharide obtained by gel chromatography on Bio-Gel P-4 of the high molecular mass material from Rt120 had a toxic effect on tumor HeLa cells but was inactive against the normal human skin fibroblast cell line. The polysaccharide from RtTA1 was inactive against either cell line. The potent inhibitory effect of the mutant DPS on tumor HeLa cells seems to be related with the differences in sugar composition.

Archives of Microbiology, 2004

Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the ... more Rhizobium leguminosarum biovar trifolii strain TA1 polysaccharide synthesis (pss) mutants in the pssD, pssP, pssT and pssO genes and altered in exopolysaccharide (EPS) synthesis were investigated. EPS-deficient mutants were also changed in lipopolysaccharide structure. All mutants exhibited varied sensitivities to detergents, ethanol and antibiotics, thus indicating changes in bacterial membrane integrity. Using pss mutants marked with the gusA gene, EPS-deficient mutants were found to have abnormalities in nodule development and to provoke severe plant defence reactions. The pss mutants that produced altered quantities of EPS with a changed degree of polymerisation generally occupied the younger developmental zones of the nodules and elicited moderate plant defence reactions.

Archiv der Pharmazie, 2012

A series of novel compounds were synthesized in reactions of N 3 -substituted amidrazones with ci... more A series of novel compounds were synthesized in reactions of N 3 -substituted amidrazones with cis-1,2cyclohexanedicarboxylic anhydride: linear, isoindole, and triazole derivatives. All new structures were confirmed by H 1 NMR and IR spectrometry as well as elemental analysis. Potential biological effects of new compounds were predicted with the Prediction of Activity Spectra for Substances (PASS) program. Antiviral, antibacterial, analgesic, and anti-inflammatory activities were experimentally verified.

BMC Microbiology

Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodule... more Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodules on roots of host plants Trifolium spp. Bacterial surface polysaccharides are crucial for establishment of a successful symbiosis with legumes that form indeterminate-type nodules, such as Trifolium, Pisum, Vicia, and Medicago spp. and aid the bacterium in withstanding osmotic and other environmental stresses. Recently, the R. leguminosarum bv. trifolii RosR regulatory protein which controls exopolysaccharide production has been identified and characterized. In this work, we extend our earlier studies to the characterization of rosR mutants which exhibit pleiotropic phenotypes. The mutants produce three times less exopolysaccharide than the wild type, and the low-molecular-weight fraction in that polymer is greatly reduced. Mutation in rosR also results in quantitative alterations in the polysaccharide constituent of lipopolysaccharide. The rosR mutants are more sensitive to surface-act...