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Chembiochem : a European Journal of... Sep 2019Lanthanide (Ln)-dependent methanol dehydrogenases (MDHs) have recently been shown to be widespread in methylotrophic bacteria. Along with the core MDH protein, XoxF,...
Lanthanide (Ln)-dependent methanol dehydrogenases (MDHs) have recently been shown to be widespread in methylotrophic bacteria. Along with the core MDH protein, XoxF, these systems contain two other proteins, XoxG (a c-type cytochrome) and XoxJ (a periplasmic binding protein of unknown function), about which little is known. In this work, we have biochemically and structurally characterized these proteins from the methyltroph Methylobacterium extorquens AM1. In contrast to results obtained in an artificial assay system, assays of XoxFs metallated with La , Ce , and Nd using their physiological electron acceptor, XoxG, display Ln-independent activities, but the K for XoxG markedly increases from La to Nd. This result suggests that XoxG's redox properties are tuned specifically for lighter Lns in XoxF, an interpretation supported by the unusually low reduction potential of XoxG (+172 mV). The X-ray crystal structure of XoxG provides a structural basis for this reduction potential and insight into the XoxG-XoxF interaction. Finally, the X-ray crystal structure of XoxJ reveals a large hydrophobic cleft and suggests a role in the activation of XoxF. These studies enrich our understanding of the underlying chemical principles that enable the activity of XoxF with multiple lanthanides in vitro and in vivo.
Topics: Alcohol Oxidoreductases; Bacterial Proteins; Cytochrome c Group; Enzyme Assays; Kinetics; Lanthanoid Series Elements; Methanol; Methylobacterium extorquens; Oxidation-Reduction; Periplasmic Binding Proteins; Rhodothermus; Saccharomyces cerevisiae
PubMed: 31017712
DOI: 10.1002/cbic.201900184 -
Applied and Environmental Microbiology Jun 2019,-Dimethylformamide (DMF) is one of the most common xenobiotic chemicals, and it can be easily emitted into the environment, where it causes harm to human beings....
,-Dimethylformamide (DMF) is one of the most common xenobiotic chemicals, and it can be easily emitted into the environment, where it causes harm to human beings. Herein, an efficient DMF-degrading strain, DM1, was isolated and identified as sp. This strain can use DMF as the sole source of carbon and nitrogen. Whole-genome sequencing of strain DM1 revealed that it has a 5.66-Mbp chromosome and a 200-kbp megaplasmid. The plasmid pLVM1 specifically harbors the genes essential for the initial steps of DMF degradation, and the chromosome carries the genes facilitating subsequent methylotrophic metabolism. Through analysis of the transcriptome sequencing data, the complete mineralization pathway and redundant gene clusters of DMF degradation were elucidated. The dimethylformamidase (DMFase) gene was heterologously expressed, and DMFase was purified and characterized. Plasmid pLVM1 is catabolically crucial for DMF utilization, as evidenced by the phenotype identification of the plasmid-free strain. This study systematically elucidates the molecular mechanisms of DMF degradation by DMF is a hazardous pollutant that has been used in the chemical industry, pharmaceutical manufacturing, and agriculture. Biodegradation as a method for removing DMF has received increasing attention. Here, we identified an efficient DMF degrader, sp. strain DM1, and characterized the complete DMF mineralization pathway and enzymatic properties of DMFase in this strain. This study provides insights into the molecular mechanisms and evolutionary advantage of DMF degradation facilitated by plasmid pLVM1 and redundant genes in strain DM1, suggesting the emergence of new ecotypes of .
Topics: Biodegradation, Environmental; Carbon; Dimethylformamide; Methylobacterium; Nitrogen; Plasmids
PubMed: 30952664
DOI: 10.1128/AEM.00275-19 -
Journal of Pediatric Hematology/oncology Jul 2020Roseomonas gilardii is a Gram-negative coccobacillus identified in immunocompromised pediatric patients. A 5-year-old male with a history of HbSβ thalassemia status... (Review)
Review
Roseomonas gilardii is a Gram-negative coccobacillus identified in immunocompromised pediatric patients. A 5-year-old male with a history of HbSβ thalassemia status postsurgical splenectomy presented to the emergency department with fever. Blood cultures grew R. gilardii at 63 hours, but the patient had been discharged home at 48 hours. The patient was readmitted for repeat cultures and initiated on meropenem for 10 days as Roseomonas spp. are often resistant to third generation cephalosporins. R. gilardii is a rare cause of bacteremia in immunocompromised patients. Clinicians should consider Roseomonas in slow growing Gram-negative rod bacteremias, and consider meropenem as empiric coverage.
Topics: Bacteremia; Child, Preschool; Gram-Negative Bacterial Infections; Humans; Male; Methylobacteriaceae; Prognosis; Thalassemia
PubMed: 30951022
DOI: 10.1097/MPH.0000000000001476 -
Planta Jun 2019Inoculation of endophytic Methylobacterium oryzae CBMB20 in salt-stressed rice plants improves photosynthesis and reduces stress volatile emissions due to mellowing of...
Inoculation of endophytic Methylobacterium oryzae CBMB20 in salt-stressed rice plants improves photosynthesis and reduces stress volatile emissions due to mellowing of ethylene-dependent responses and activating vacuolar H-ATPase. The objective of this study was to analyze the impact of ACC (1-aminocyclopropane-1-carboxylate) deaminase-producing Methylobacterium oryzae CBMB20 in acclimation of plant to salt stress by controlling photosynthetic characteristics and volatile emission in salt-sensitive (IR29) and moderately salt-resistant (FL478) rice (Oryza sativa L.) cultivars. Saline levels of 50 mM and 100 mM NaCl with and without bacteria inoculation were applied, and the temporal changes in stress response and salinity resistance were assessed by monitoring photosynthetic characteristics, ACC accumulation, ACC oxidase activity (ACO), vacuolar H ATPase activity, and volatile organic compound (VOC) emissions. Salt stress considerably reduced photosynthetic rate, stomatal conductance, PSII efficiency and vacuolar H ATPase activity, but it increased ACC accumulation, ACO activity, green leaf volatiles, mono- and sesquiterpenes, and other stress volatiles. These responses were enhanced with increasing salt stress and time. However, rice cultivars treated with CBMB20 showed improved plant vacuolar H ATPase activity, photosynthetic characteristics and decreased ACC accumulation, ACO activity and VOC emission. The bacteria-dependent changes were greater in the IR29 cultivar. These results indicate that decreasing photosynthesis and vacuolar H ATPase activity rates and increasing VOC emission rates in response to high-salinity stress were effectively mitigated by M. oryzae CBMB20 inoculation.
Topics: Amino Acid Oxidoreductases; Carbon-Carbon Lyases; Endophytes; Ethylenes; Genotype; Methylobacterium; Oryza; Phenotype; Photosynthesis; Plant Growth Regulators; Plant Proteins; Salinity; Salt Stress; Stress, Physiological; Vacuolar Proton-Translocating ATPases; Volatile Organic Compounds
PubMed: 30877435
DOI: 10.1007/s00425-019-03139-w -
Scientific Reports Mar 2019Lanthanide (Ln) elements are utilized as cofactors for catalysis by XoxF-type methanol dehydrogenases (MDHs). A primary assumption is that XoxF enzymes produce formate...
Lanthanide (Ln) elements are utilized as cofactors for catalysis by XoxF-type methanol dehydrogenases (MDHs). A primary assumption is that XoxF enzymes produce formate from methanol oxidation, which could impact organisms that require formaldehyde for assimilation. We report genetic and phenotypic evidence showing that XoxF1 (MexAM1_1740) from Methylobacterium extorquens AM1 produces formaldehyde, and not formate, during growth with methanol. Enzyme purified with lanthanum or neodymium oxidizes formaldehyde. However, formaldehyde oxidation via 2,6-dichlorophenol-indophenol (DCPIP) reduction is not detected in cell-free extracts from wild-type strain methanol- and lanthanum-grown cultures. Formaldehyde activating enzyme (Fae) is required for Ln methylotrophic growth, demonstrating that XoxF1-mediated production of formaldehyde is essential. Addition of exogenous lanthanum increases growth rate with methanol by 9-12% but does not correlate with changes to methanol consumption or formaldehyde accumulation. Transcriptomics analysis of lanthanum methanol growth shows upregulation of xox1 and downregulation of mxa genes, consistent with the Ln-switch, no differential expression of formaldehyde conversion genes, downregulation of pyrroloquinoline quinone (PQQ) biosynthesis genes, and upregulation of fdh4 formate dehydrogenase (FDH) genes. Additionally, the Ln-dependent ethanol dehydrogenase ExaF reduces methanol sensitivity in the fae mutant strain when lanthanides are present, providing evidence for the capacity of an auxiliary role for ExaF during Ln-dependent methylotrophy.
Topics: Alcohol Oxidoreductases; Bacterial Proteins; Biocatalysis; Biosynthetic Pathways; Coenzymes; Enzyme Assays; Formaldehyde; Formate Dehydrogenases; Gene Expression Profiling; Genes, Bacterial; Lanthanoid Series Elements; Methanol; Methylobacterium extorquens; Oxidation-Reduction
PubMed: 30862918
DOI: 10.1038/s41598-019-41043-1 -
Journal of the American Chemical Society Mar 2019Understanding the biosynthesis of cofactors is fundamental to the life sciences, yet to date a few important pathways remain unresolved. One example is the redox...
Understanding the biosynthesis of cofactors is fundamental to the life sciences, yet to date a few important pathways remain unresolved. One example is the redox cofactor pyrroloquinoline quinone (PQQ), which is critical for C1 metabolism in many microorganisms, a disproportionate number of which are opportunistic human pathogens. While the initial and final steps of PQQ biosynthesis, involving PqqD/E and PqqC, have been elucidated, the precise nature and order of the remaining transformations in the pathway are unknown. Here we show evidence that the remaining essential biosynthetic enzyme PqqB is an iron-dependent hydroxylase catalyzing oxygen-insertion reactions that are proposed to produce the quinone moiety of the mature PQQ cofactor. The demonstrated reactions of PqqB are unprecedented within the metallo β-lactamase protein family and expand the catalytic repertoire of nonheme iron hydroxylases. These new findings also generate a nearly complete description of the PQQ biosynthetic pathway.
Topics: Bacterial Proteins; Catalysis; Dihydroxyphenylalanine; Hydroxylation; Iron; Methylobacterium extorquens; Mixed Function Oxygenases; Models, Chemical; Zinc
PubMed: 30811189
DOI: 10.1021/jacs.8b13453 -
BMC Infectious Diseases Feb 2019Roseomonas mucosa, as a Gram-negative coccobacilli, is an opportunistic pathogen that has rarely been reported in human infections. Here we describe a case of bacteremia... (Review)
Review
BACKGROUND
Roseomonas mucosa, as a Gram-negative coccobacilli, is an opportunistic pathogen that has rarely been reported in human infections. Here we describe a case of bacteremia in an infective endocarditis patient with systemic lupus erythematosus (SLE).
CASE PRESENTATIONS
A 44-year-old female patient with SLE suffered bacteremia caused by Roseomonas mucosa complicated with infective endocarditis (IE). The patient started on treatment with piperacillin-tazobactam and levofloxacin against Roseomonas mucosa, which was switched after 4 days to meropenem and amikacin for an additional 2 weeks. She had a favorable outcome with a 6-week course of intravenous antibiotic therapy.
DISCUSSION AND CONCLUSIONS
Roseomonas mucosa is rarely reported in IE patients; therefore, we report the case in order to improve our ability to identify this pathogen and expand the range of known bacterial causes of infective endocarditis.
Topics: Adult; Amikacin; Anti-Bacterial Agents; Bacteremia; Endocarditis; Endocarditis, Bacterial; Female; Gram-Negative Bacterial Infections; Humans; Levofloxacin; Lupus Erythematosus, Systemic; Methylobacteriaceae; Piperacillin, Tazobactam Drug Combination
PubMed: 30755159
DOI: 10.1186/s12879-019-3774-0 -
Frontiers in Cellular and Infection... 2018As therapies for atopic dermatitis (AD) based on live biotherapeutic products (LBP) are developed, the potential displacement of biotherapeutic strains, and species to...
As therapies for atopic dermatitis (AD) based on live biotherapeutic products (LBP) are developed, the potential displacement of biotherapeutic strains, and species to mucosal sites where they are not naturally found is of investigative interest. However, formal assessment of the toxicity potential of healthy skin commensal organisms has not been reported in the literature. Our previous research indicates that topical application of live to treat AD was associated with clinical benefit on the skin, but the effects of exposure via inhalation, eye inoculation, and ingestion were unknown. Herein we report our findings from mice inoculated with commensal strains of , coagulase negative (CNS), and . Bacterial isolates were collected under clinical trial NCT03018275, however these results do not represent an interventional clinical trial. Our tested R. mucosa isolates did not display significant infection or inflammation. However, neutropenic mice inoculated with CNS had infection without major inflammation in pulmonary models. In contrast, systemic infection generated hepatic and splenic pathology for and CNS, which was worsened by the presence of neutropenia. Our results suggest that LBP derived from bacteria without significant infectivity histories, such as , may represent safer options than known pathobionts like and spp. Overall, these results suggest that topically applied LBP from select skin commensals are likely to present safe therapeutic options and reinforce our prior clinical findings.
Topics: Animals; Bacterial Infections; Carrier State; Disease Models, Animal; Methylobacteriaceae; Mice; Probiotics; Pseudomonas aeruginosa; Staphylococcus; Symbiosis; Virulence
PubMed: 30719426
DOI: 10.3389/fcimb.2018.00451 -
Molecular Microbiology May 2019Until recently, rare-earth elements (REEs) had been thought to be biologically inactive. This view changed with the discovery of the methanol dehydrogenase XoxF that...
Until recently, rare-earth elements (REEs) had been thought to be biologically inactive. This view changed with the discovery of the methanol dehydrogenase XoxF that strictly relies on REEs for its activity. Some methylotrophs only contain xoxF, while others, including the model phyllosphere colonizer Methylobacterium extorquens PA1, harbor this gene in addition to mxaFI encoding a Ca -dependent enzyme. Here we found that REEs induce the expression of xoxF in M. extorquens PA1, while repressing mxaFI, suggesting that XoxF is the preferred methanol dehydrogenase in the presence of sufficient amounts of REE. Using reporter assays and a suppressor screen, we found that lanthanum (La ) is sensed both in a XoxF-dependent and independent manner. Furthermore, we investigated the role of REEs during Arabidopsis thaliana colonization. Element analysis of the phyllosphere revealed the presence of several REEs at concentrations up to 10 μg per g dry weight. Complementary proteome analyses of M. extorquens PA1 identified XoxF as a top induced protein in planta and a core set of La -regulated proteins under defined artificial media conditions. Among these was a REE-binding protein that is encoded next to a gene for a TonB-dependent transporter. The latter was essential for REE-dependent growth on methanol indicating chelator-assisted uptake of REEs.
Topics: Alcohol Oxidoreductases; Arabidopsis; Gene Expression Regulation, Bacterial; Lanthanum; Methanol; Methylobacterium extorquens; Proteome
PubMed: 30653750
DOI: 10.1111/mmi.14208 -
Pathogens (Basel, Switzerland) Jan 2019The bacterial biota in larvae of , a serious pest of cultivated stone-fruit trees in the West Palearctic, was revealed for the first time using the MiSeq platform. The...
The bacterial biota in larvae of , a serious pest of cultivated stone-fruit trees in the West Palearctic, was revealed for the first time using the MiSeq platform. The core bacterial community remained the same in neonates whether upon hatching or grown on peach plants or an artificial diet, suggesting that larvae acquire much of their bacterial biome from the parent adult. Reads affiliated with class levels and (phylum ca. 86%), and (ca. 14%) were highly abundant. Most diverse reads belong to the families (50%), (20%), (9%), (7%) and (4.5%). About two-thirds of the reads are affiliated with the genera , , , and , which encompass species displaying cellulolytic and lipolytic activities. This study provides a foundation for future studies to elucidate the roles of bacterial biota in .
PubMed: 30621355
DOI: 10.3390/pathogens8010004