scholarly journals The Role of Prophage for Genome Diversification within a Clonal Lineage of Lactobacillus johnsonii: Characterization of the Defective Prophage LJ771

2008 ◽  
Vol 190 (17) ◽  
pp. 5806-5813 ◽  
Author(s):  
Emmanuel Denou ◽  
Raymond David Pridmore ◽  
Marco Ventura ◽  
Anne-Cécile Pittet ◽  
Marie-Camille Zwahlen ◽  
...  
Keyword(s):  
Methionine Sulfoxide ◽  
Gene Cassette ◽  
Methionine Sulfoxide Reductase ◽  
Toxin Gene ◽  
Clonal Lineage ◽  
Lactobacillus Johnsonii ◽  
Reductase Gene ◽  
Phage Dna

ABSTRACT Two independent isolates of the gut commensal Lactobacillus johnsonii were sequenced. These isolates belonged to the same clonal lineage and differed mainly by a 40.8-kb prophage, LJ771, belonging to the Sfi11 phage lineage. LJ771 shares close DNA sequence identity with Lactobacillus gasseri prophages. LJ771 coexists as an integrated prophage and excised circular phage DNA, but phage DNA packaged into extracellular phage particles was not detected. Between the phage lysin gene and attR a likely mazE (“antitoxin”)/pemK (“toxin”) gene cassette was detected in LJ771 but not in the L. gasseri prophages. Expressed pemK could be cloned in Escherichia coli only together with the mazE gene. LJ771 was shown to be highly stable and could be cured only by coexpression of mazE from a plasmid. The prophage was integrated into the methionine sulfoxide reductase gene (msrA) and complemented the 5′ end of this gene, creating a protein with a slightly altered N-terminal sequence. The two L. johnsonii strains had identical in vitro growth and in vivo gut persistence phenotypes. Also, in an isogenic background, the presence of the prophage resulted in no growth disadvantage.

scholarly journals Identification of Lactobacillus reuteri Genes Specifically Induced in the Mouse Gastrointestinal Tract

2003 ◽  
Vol 69 (4) ◽  
pp. 2044-2051 ◽  
Author(s):  
Jens Walter ◽  
Nicholas C. K. Heng ◽  
Walter P. Hammes ◽  
Diane M. Loach ◽  
Gerald W. Tannock ◽  
...  
Keyword(s):  
Gastrointestinal Tract ◽  
Reporter Gene ◽  
Stress Responses ◽  
Lactobacillus Reuteri ◽  
Xylose Isomerase ◽  
Methionine Sulfoxide ◽  
Methionine Sulfoxide Reductase ◽  
Chromosomal Dna

ABSTRACT Lactobacilli are common inhabitants of the gastrointestinal tracts of mammals and have received considerable attention due to their putative health-promoting properties. Little is known about the traits that enhance the ability of these bacteria to inhabit the gastrointestinal tract. In this paper we describe the development and application of a strategy based on in vivo expression technology (IVET) that enables detection of Lactobacillus reuteri genes specifically induced in the murine gut. A plasmid-based system was constructed containing ′ermGT (which confers lincomycin resistance) as the primary reporter gene for selection of promoters active in the gastrointestinal tract of mice treated with lincomycin. A second reporter gene, ′bglM (β-glucanase), allowed differentiation between constitutive and in vivo inducible promoters. The system was successfully tested in vitro and in vivo by using a constitutive promoter. Application of the IVET system with chromosomal DNA of L. reuteri 100-23 and reconstituted lactobacillus-free mice revealed three genes induced specifically during colonization. Two of the sequences showed homology to genes encoding xylose isomerase (xylA) and peptide methionine sulfoxide reductase (msrB), which are involved in nutrient acquisition and stress responses, respectively. The third locus showed homology to the gene encoding a protein whose function is not known. Our IVET system has the potential to identify genes of lactobacilli that have not previously been functionally characterized but which may be essential for growth of these bacteria in the gastrointestinal ecosystem.

scholarly journals Methionine sulfoxide reductase 2 reversibly regulates Mge1, a cochaperone of mitochondrial Hsp70, during oxidative stress

2015 ◽  
Vol 26 (3) ◽  
pp. 406-419 ◽  
Author(s):  
Praveen Kumar Allu ◽  
Adinarayana Marada ◽  
Yerranna Boggula ◽  
Srinivasu Karri ◽  
Thanuja Krishnamoorthy ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Methionine Sulfoxide ◽  
Methionine Sulfoxide Reductase ◽  
Yeast Cells ◽  
Nucleotide Exchange ◽  
New Paradigm ◽  
Methionine Sulfoxide Reductases ◽  
Mitochondrial Hsp70

Peptide methionine sulfoxide reductases are conserved enzymes that reduce oxidized methionines in protein(s). Although these reductases have been implicated in several human diseases, there is a dearth of information on the identity of their physiological substrates. By using Saccharomyces cerevisiae as a model, we show that of the two methionine sulfoxide reductases (MXR1, MXR2), deletion of mitochondrial MXR2 renders yeast cells more sensitive to oxidative stress than the cytosolic MXR1. Our earlier studies showed that Mge1, an evolutionarily conserved nucleotide exchange factor of Hsp70, acts as an oxidative sensor to regulate mitochondrial Hsp70. In the present study, we show that Mxr2 regulates Mge1 by selectively reducing MetO at position 155 and restores the activity of Mge1 both in vitro and in vivo. Mge1 M155L mutant rescues the slow-growth phenotype and aggregation of proteins of mxr2Δ strain during oxidative stress. By identifying the first mitochondrial substrate for Mxrs, we add a new paradigm to the regulation of the oxidative stress response pathway.

scholarly journals Expression of magA in Legionella pneumophila Philadelphia-1 Is Developmentally Regulated and a Marker of Formation of Mature Intracellular Forms

2004 ◽  
Vol 186 (10) ◽  
pp. 3038-3045 ◽  
Author(s):  
Margot F. Hiltz ◽  
Gary R. Sisson ◽  
Ann Karen C. Brassinga ◽  
Elizabeth Garduno ◽  
Rafael A. Garduno ◽  
...  
Keyword(s):  
Hela Cells ◽  
Legionella Pneumophila ◽  
Protein Profile ◽  
Methionine Sulfoxide ◽  
Growth Cycle ◽  
Specific Protein ◽  
Methionine Sulfoxide Reductase ◽  
Developmental Cycle

ABSTRACT Legionella pneumophila displays a biphasic developmental cycle in which replicating forms (RFs) differentiate postexponentially into highly infectious, cyst-like mature intracellular forms (MIFs). Using comparative protein profile analyses (MIFs versus RFs), we identified a 20-kDa protein, previously annotated as “Mip-like” protein, that was enriched in MIFs. However, this 20-kDa protein shared no similarity with Mip, a well-characterized peptidyl-prolyl isomerase of L. pneumophila, and for clarity we renamed it MagA (for “MIF-associated gene”). We monitored MagA levels across the growth cycle (in vitro and in vivo) by immunoblotting and established that MagA levels increased postexponentially in vitro (∼3-fold) and nearly 10-fold during MIF morphogenesis in HeLa cells. DNA sequence analysis of the magA locus revealed an upstream divergently transcribed gene, msrA, encoding a peptide methionine sulfoxide reductase and a shared promoter region containing direct and indirect repeat sequences as well as −10 hexamers often associated with stationary-phase regulation. While MagA has no known function, it contains a conserved CXXC motif commonly found in members of the thioredoxin reductase family and in AhpD reductases that are associated with alkylhydroperoxide reductase (AhpC), suggesting a possible role in protection from oxidative stress. MIFs from L. pneumophila strain Lp02 containing a magA deletion exhibited differences in Giménez staining, as well as an apparent increase in cytopathology to HeLa cells, but otherwise were unaltered in virulence traits. As demonstrated by this study, MagA appears to be a MIF-specific protein expressed late in intracellular growth that may serve as a useful marker of development.

scholarly journals In Vitro and In Vivo Oxidation of Methionine Residues in Small, Acid-Soluble Spore Proteins fromBacillus Species

1998 ◽  
Vol 180 (10) ◽  
pp. 2694-2700 ◽  
Author(s):  
Christopher S. Hayes ◽  
Berenice Illades-Aguiar ◽  
Lilliam Casillas-Martinez ◽  
Peter Setlow
Keyword(s):  
Dna Binding ◽  
Methionine Sulfoxide ◽  
Small Degree ◽  
Methionine Sulfoxide Reductase ◽  
Mutant Form ◽  
Wild Type ◽  
Spore Proteins ◽  
Methionine Residues

ABSTRACT Methionine residues in α/β-type small, acid-soluble spore proteins (SASP) of Bacillus species were readily oxidized to methionine sulfoxide in vitro by t-butyl hydroperoxide (tBHP) or hydrogen peroxide (H2O2). These oxidized α/β-type SASP no longer bound to DNA effectively, but DNA binding protected α/β-type SASP against methionine oxidation by peroxides in vitro. Incubation of an oxidized α/β-type SASP with peptidyl methionine sulfoxide reductase (MsrA), which can reduce methionine sulfoxide residues back to methionine, restored the α/β-type SASP’s ability to bind to DNA. Both tBHP and H2O2 caused some oxidation of the two methionine residues of an α/β-type SASP (SspC) in spores ofBacillus subtilis, although one methionine which is highly conserved in α/β-type SASP was only oxidized to a small degree. However, much more methionine sulfoxide was generated by peroxide treatment of spores carrying a mutant form of SspC which has a lower affinity for DNA. MsrA activity was present in wild-type B. subtilis spores. However, msrA mutant spores were no more sensitive to H2O2 than were wild-type spores. The major mechanism operating for dealing with oxidative damage to α/β-type SASP in spores is DNA binding, which protects the protein’s methionine residues from oxidation both in vitro and in vivo. This may be important in vivo since α/β-type SASP containing oxidized methionine residues no longer bind DNA well and α/β-type SASP-DNA binding is essential for long-term spore survival.

scholarly journals Methionine Sulfoxide Reductase B Regulates the Activity of Ascorbate Peroxidase of Banana Fruit

Antioxidants ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 310
Author(s):  
Lu Xiao ◽  
Guoxiang Jiang ◽  
Huiling Yan ◽  
Hongmei Lai ◽  
Xinguo Su ◽  
...  
Keyword(s):  
Ascorbate Peroxidase ◽  
Posttranslational Modifications ◽  
Stress Responses ◽  
Redox State ◽  
Methionine Sulfoxide ◽  
Methionine Sulfoxide Reductase ◽  
Banana Fruit ◽  
Compound I

Ascorbate peroxidase (APX) is a key antioxidant enzyme that is involved in diverse developmental and physiological process and stress responses by scavenging H2O2 in plants. APX itself is also subjected to multiple posttranslational modifications (PTMs). However, redox-mediated PTM of APX in plants remains poorly understood. Here, we identified and confirmed that MaAPX1 interacts with methionine sulfoxide reductase B2 (MsrB2) in bananas. Ectopic overexpression of MaAPX1 delays the detached leaf senescence induced by darkness in Arabidopsis. Sulfoxidation of MaAPX1, i.e., methionine oxidation, leads to loss of the activity, which is repaired partially by MaMsrB2. Moreover, mimicking sulfoxidation by mutating Met36 to Gln also decreases its activity in vitro and in vivo, whereas substitution of Met36 with Val36 to mimic the blocking of sulfoxidation has little effect on APX activity. Spectral analysis showed that mimicking sulfoxidation of Met36 hinders the formation of compound I, the first intermediate between APX and H2O2. Our findings demonstrate that the redox state of methionine in MaAPX1 is critical to its activity, and MaMsrB2 can regulate the redox state and activity of MaAPX1. Our results revealed a novel post-translational redox modification of APX.

scholarly journals An NMR-based biosensor to measure stereo-specific methionine sulfoxide reductase (MSR) activities in vitro and in vivo

2020 ◽  
Author(s):  
Carolina Sánchez-López ◽  
Natalia Labadie ◽  
Verónica A. Lombardo ◽  
Franco A. Biglione ◽  
Bruno Manta ◽  
...  
Keyword(s):  
Methionine Sulfoxide ◽  
Methionine Sulfoxide Reductase ◽  
Substrate Specificities ◽  
Age Related ◽  
Native Cell ◽  
Highly Sensitive ◽  
Sensitive Tool ◽  
Single Reaction

AbstractOxidation of protein methionines to methionine-sulfoxides (MetOx) is associated with several age-related diseases. In healthy cells, MetOx is reduced to methionine by two families of conserved methionine sulfoxide reductase enzymes, MSRA and MSRB that specifically target the S- or R-diastereoisomers of methionine-sulfoxides, respectively. To directly interrogate MSRA and MSRB functions in cellular settings, we developed an NMR-based biosensor that we call CarMetOx to simultaneously measure both enzyme activities in single reaction setups. We demonstrate the suitability of our strategy to delineate MSR functions in complex biological environments that range from native cell lysates to zebrafish embryos. Thereby, we establish differences in substrate specificities between prokaryotic and eukaryotic MSRs and introduce CarMetOx as a highly sensitive tool for studying therapeutic targets of oxidative stress-related human diseases and redox regulated signaling pathways. Our approach further extends high-resolution in-cell NMR measurements of exogenously delivered biomolecules to an entire multicellular organism.

scholarly journals An NMR‐Based Biosensor to Measure Stereospecific Methionine Sulfoxide Reductase Activities in Vitro and in Vivo**

2020 ◽  
Vol 26 (65) ◽  
pp. 14838-14843
Author(s):  
Carolina Sánchez‐López ◽  
Natalia Labadie ◽  
Verónica A. Lombardo ◽  
Franco A. Biglione ◽  
Bruno Manta ◽  
...  
Keyword(s):  
Methionine Sulfoxide ◽  
Methionine Sulfoxide Reductase

scholarly journals Molecular Basis of In Vivo Resistance to Sulfadoxine-Pyrimethamine in African Adult Patients Infected withPlasmodium falciparum Malaria Parasites

1998 ◽  
Vol 42 (7) ◽  
pp. 1811-1814 ◽  
Author(s):  
Leonardo K. Basco ◽  
Rachida Tahar ◽  
Pascal Ringwald
Keyword(s):  
Dihydrofolate Reductase ◽  
Point Mutations ◽  
Gene Mutations ◽  
Adult Patients ◽  
Wild Type ◽  
Dihydropteroate Synthase ◽  
Reductase Gene ◽  
Parasite Populations

ABSTRACT In vitro sulfadoxine and pyrimethamine resistance has been associated with point mutations in the dihydropteroate synthase and dihydrofolate reductase domains, respectively, but the in vivo relevance of these point mutations has not been well established. To analyze the correlation between genotype and phenotype, 10 Cameroonian adult patients were treated with sulfadoxine-pyrimethamine and followed up for 28 days. After losses to follow-up (n = 1) or elimination of DNA samples due to mixed parasite populations with pyrimethamine-sensitive and pyrimethamine-resistant profiles (n = 3), parasite genomic DNA from day 0 blood samples of six patients were analyzed by DNA sequencing. Three patients who were cured had isolates characterized by a wild-type or mutant dihydrofolate reductase gene (with one or two mutations) and a wild-type dihydropteroate synthase gene. Three other patients who failed to respond to sulfadoxine-pyrimethamine treatment carried isolates with triple dihydrofolate reductase gene mutations and either a wild-type or a mutant dihydropteroate synthase gene. Three dihydrofolate reductase gene codons (51, 59, and 108) may be reliable genetic markers that can accurately predict the clinical outcome of sulfadoxine-pyrimethamine treatment in Africa.

Channeling phage DNA through membranes: from in vivo to in vitro

2003 ◽  
Vol 154 (4) ◽  
pp. 283-287 ◽  
Author(s):  
Lucienne Letellier ◽  
Pascale Boulanger ◽  
Marta de Frutos ◽  
Pierre Jacquot
Keyword(s):  
Phage Dna

scholarly journals Lactobacillus johnsonii La1 Attenuates Helicobacter pylori-Associated Gastritis and Reduces Levels of Proinflammatory Chemokines in C57BL/6 Mice

2005 ◽  
Vol 12 (12) ◽  
pp. 1378-1386 ◽  
Author(s):  
Dionyssios N. Sgouras ◽  
Effrosini G. Panayotopoulou ◽  
Beatriz Martinez-Gonzalez ◽  
Kalliopi Petraki ◽  
Spyros Michopoulos ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Lamina Propria ◽  
Serum Levels ◽  
Favorable Effect ◽  
Lactobacillus Johnsonii ◽  
Ags Cells ◽  
Inflammatory Infiltration ◽  
H Pylori

ABSTRACT In clinical settings, Lactobacillus johnsonii La1 administration has been reported to have a favorable effect on Helicobacter pylori-associated gastritis, although the mechanism remains unclear. We administered, continuously through the water supply, live La1 to H. pylori-infected C57BL/6 mice and followed colonization, the development of H. pylori-associated gastritis in the lamina propria, and the levels of proinflammatory chemokines macrophage inflammatory protein 2 (MIP-2) and keratinocyte-derived cytokine (KC) in the serum and gastric tissue over a period of 3 months. We documented a significant attenuation in both lymphocytic (P = 0.038) and neutrophilic (P = 0.003) inflammatory infiltration in the lamina propria as well as in the circulating levels of anti-H. pylori immunoglobulin G antibodies (P = 0.003), although we did not observe a suppressive effect of La1 on H. pylori colonizing numbers. Other lactobacilli, such as L. amylovorus DCE 471 and L. acidophilus IBB 801, did not attenuate H. pylori-associated gastritis to the same extent. MIP-2 serum levels were distinctly reduced during the early stages of H. pylori infection in the La1-treated animals, as were gastric mucosal levels of MIP-2 and KC. Finally, we also observed a significant reduction (P = 0.046) in H. pylori-induced interleukin-8 secretion by human adenocarcinoma AGS cells in vitro in the presence of neutralized (pH 6.8) La1 spent culture supernatants, without concomitant loss of H. pylori viability. These observations suggest that during the early infection stages, administration of La1 can attenuate H. pylori-induced gastritis in vivo, possibly by reducing proinflammatory chemotactic signals responsible for the recruitment of lymphocytes and neutrophils in the lamina propria.

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