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plasmids of carotenoid-producing paracoccus spp. (alphaproteobacteria) - structure, diversity and evolution.plasmids are components of many bacterial genomes. they enable the spread of a large pool of genetic information via lateral gene transfer. many bacterial strains contain mega-sized replicons and these are particularly common in alphaproteobacteria. considerably less is known about smaller alphaproteobacterial plasmids. we analyzed the genomes of 14 such plasmids residing in 4 multireplicon carotenoid-producing strains of the genus paracoccus (alphaproteobacteria): p. aestuarii dsm 19484, p. hae ...201324260361
fluorescence-based bacterial bioreporter for specific detection of methyl halide emissions in the environment.methyl halides are volatile one-carbon compounds responsible for substantial depletion of stratospheric ozone. among them, chloromethane (ch3cl) is the most abundant halogenated hydrocarbon in the atmosphere. global budgets of methyl halides in the environment are still poorly understood due to uncertainties in their natural sources, mainly from vegetation, and their sinks, which include chloromethane-degrading bacteria. a bacterial bioreporter for the detection of methyl halides was developed o ...201323956392
metabolic adaptation and trophic strategies of soil bacteria-c1- metabolism and sulfur chemolithotrophy in starkeya novella.the highly diverse and metabolically versatile microbial communities found in soil environments are major contributors to the global carbon, nitrogen, and sulfur cycles. we have used a combination of genome -based pathway analysis with proteomics and gene expression studies to investigate metabolic adaptation in a representative of these bacteria, starkeya novella, which was originally isolated from agricultural soil. this bacterium was the first facultative sulfur chemolithoautotroph that was i ...201324146664
an oxidoreductase from 'alphonso' mango catalyzing biosynthesis of furaneol and reduction of reactive carbonyls.two furanones, furaneol (4-hydroxy-2,5-dimethyl-3(2h)-furanone) and mesifuran (2,5-dimethyl-4-methoxy-3(2h)-furanone), are important constituents of flavor of the alphonso cultivar of mango (mangifera indica). to get insights into the biosynthesis of these furanones, we isolated an enone oxidoreductase gene from the alphonso mango. it has high sequence similarity to an alkenal/one oxidoreductase from cucumber (79% identity) and enone oxidoreductases from tomato (73% identity) and strawberry (72% ...201324133645
hydrogen and carbon isotope fractionation during degradation of chloromethane by methylotrophic bacteria.chloromethane (ch3 cl) is a widely studied volatile halocarbon involved in the destruction of ozone in the stratosphere. nevertheless, its global budget still remains debated. stable isotope analysis is a powerful tool to constrain fluxes of chloromethane between various environmental compartments which involve a multiplicity of sources and sinks, and both biotic and abiotic processes. in this study, we measured hydrogen and carbon isotope fractionation of the remaining untransformed chlorometha ...201324019296
autoinhibition and signaling by the switch ii motif in the g-protein chaperone of a radical b12 enzyme.meab is an accessory gtpase protein involved in the assembly, protection, and reactivation of 5'-deoxyadenosyl cobalamin-dependent methylmalonyl-coa mutase (mcm). mutations in the human ortholog of meab result in methylmalonic aciduria, an inborn error of metabolism. g-proteins typically utilize conserved switch i and ii motifs for signaling to effector proteins via conformational changes elicited by nucleotide binding and hydrolysis. our recent discovery that meab utilizes an unusual switch iii ...201323996001
the (d)evolution of methanotrophy in the beijerinckiaceae--a comparative genomics analysis.the alphaproteobacterial family beijerinckiaceae contains generalists that grow on a wide range of substrates, and specialists that grow only on methane and methanol. we investigated the evolution of this family by comparing the genomes of the generalist organotroph beijerinckia indica, the facultative methanotroph methylocella silvestris and the obligate methanotroph methylocapsa acidiphila. highly resolved phylogenetic construction based on universally conserved genes demonstrated that the bei ...201323985741
the (d)evolution of methanotrophy in the beijerinckiaceae--a comparative genomics analysis.the alphaproteobacterial family beijerinckiaceae contains generalists that grow on a wide range of substrates, and specialists that grow only on methane and methanol. we investigated the evolution of this family by comparing the genomes of the generalist organotroph beijerinckia indica, the facultative methanotroph methylocella silvestris and the obligate methanotroph methylocapsa acidiphila. highly resolved phylogenetic construction based on universally conserved genes demonstrated that the bei ...201323985741
isolation of optically targeted single bacteria by application of fluidic force microscopy to aerobic anoxygenic phototrophs from the phyllosphere.in their natural environment, bacteria often behave differently than they do under laboratory conditions. to gain insight into the physiology of bacteria in situ, dedicated approaches are required to monitor their adaptations and specific behaviors under environmental conditions. optical microscopy is crucial for the observation of fundamental characteristics of bacteria, such as cell shape, size, and marker gene expression. here, fluidic force microscopy (fluidfm) was exploited to isolate optic ...201323770907
mercurial-resistance determinants in pseudomonas strain k-62 plasmid pmr68.we report the complete nucleotide sequence of plasmid pmr68, isolated from pseudomonas strain k-62, two plasmids contribute to broad-spectrum mercury resistance and that the mer operon from one of them (pmr26) has been previously characterized. the plasmid was 71,020 bp in length and contained 75 coding regions. three mer gene clusters were identified. the first comprised merr-orf4-orf5-mert1-merp1-merf-mera-merb1, which confers bacterial resistance to mercuric ions and organomercury. the second ...201323890172
function and x-ray crystal structure of escherichia coli yfde.many food plants accumulate oxalate, which humans absorb but do not metabolize, leading to the formation of urinary stones. the commensal bacterium oxalobacter formigenes consumes oxalate by converting it to oxalyl-coa, which is decarboxylated by oxalyl-coa decarboxylase (oxc). oxc and the class iii coa-transferase formyl-coa:oxalate coa-transferase (fcoct) are widespread among bacteria, including many that have no apparent ability to degrade or to resist external oxalate. the evga acid response ...201323935849
a switch iii motif relays signaling between a b12 enzyme and its g-protein chaperone.fidelity during cofactor assembly is essential for the proper functioning of metalloenzymes and is ensured by specific chaperones. meab, a g-protein chaperone for the coenzyme b12-dependent radical enzyme methylmalonyl-coa mutase (mcm), uses the energy of gtp binding, hydrolysis or both to regulate cofactor loading into mcm, protect mcm from inactivation and rescue mcm that is inactivated during turnover. typically, g proteins signal to client proteins using the conformationally mobile switch i ...201323873214
blue light regulated two-component systems: enzymatic and functional analyses of light-oxygen-voltage (lov)-histidine kinases and downstream response regulators.light is an essential environmental cue for diverse organisms. many prokaryotic blue light photoreceptors use light, oxygen, voltage (lov) sensory domains to control the activities of diverse output domains, including histidine kinases (hk). upon activation, these proteins autophosphorylate a histidine residue before subsequently transferring the phosphate to an aspartate residue in the receiver domain of a cognate response regulator (rr). such phosphorylation activates the output domain of the ...201323806044
multistep, eight-electron oxidation catalyzed by the cofactorless oxidase, pqqc: identification of chemical intermediates and their dependence on molecular oxygen.the final step of the biosynthesis of prokaryotic cofactor pqq is catalyzed by pqqc, a cofactorless oxidase that brings about a ring closure and overall eight-electron oxidation of its substrate. time-dependent acid quenching and subsequent high-performance liquid chromatography separation and mass spectrometric analyses of reaction mixtures were performed to correlate the structures of intermediates with previously observed uv-visible signatures. the reaction is composed of four stepwise oxidat ...201323718207
draft genome sequence of methylobacterium mesophilicum strain sr1.6/6, isolated from citrus sinensis.methylobacterium mesophilicum strain sr1.6/6 is an endophytic bacterium isolated from a surface-sterilized citrus sinensis branch. ecological and biotechnological aspects of this bacterium, such as the genes involved in its association with the host plant and the primary oxidation of methanol, were annotated in the draft genome.201323788544
identification of opportunistic pathogenic bacteria in drinking water samples of different rural health centers and their clinical impacts on humans.international drinking water quality monitoring programs have been established in order to prevent or to reduce the risk of contracting water-related infections. a survey was performed on groundwater-derived drinking water from 13 different hospitals in the mahabubnagar district. a total of 55 bacterial strains were isolated which belonged to both gram-positive and gram-negative bacteria. all the taxa were identified based on the 16s rrna gene sequence analysis based on which they are phylogenet ...201323862144
development of an optimized medium, strain and high-throughput culturing methods for methylobacterium extorquens.methylobacterium extorquens strains are the best-studied methylotrophic model system, and their metabolism of single carbon compounds has been studied for over 50 years. here we develop a new system for high-throughput batch culture of m. extorquens in microtiter plates by jointly optimizing the properties of the organism, the growth media and the culturing system. after removing cellulose synthase genes in m. extorquens strains am1 and pa1 to prevent biofilm formation, we found that currently a ...201323646164
global molecular analyses of methane metabolism in methanotrophic alphaproteobacterium, methylosinus trichosporium ob3b. part ii. metabolomics and 13c-labeling study.in this work we use metabolomics and (13)c-labeling data to refine central metabolic pathways for methane utilization in methylosinus trichosporium ob3b, a model alphaproteobacterial methanotrophic bacterium. we demonstrate here that similar to non-methane utilizing methylotrophic alphaproteobacteria the core metabolism of the microbe is represented by several tightly connected metabolic cycles, such as the serine pathway, the ethylmalonyl-coa (emc) pathway, and the citric acid (tca) cycle. both ...201323565113
global molecular analyses of methane metabolism in methanotrophic alphaproteobacterium, methylosinus trichosporium ob3b. part i: transcriptomic study.methane utilizing bacteria (methanotrophs) are important in both environmental and biotechnological applications, due to their ability to convert methane to multicarbon compounds. however, systems-level studies of methane metabolism have not been carried out in methanotrophs. in this work we have integrated genomic and transcriptomic information to provide an overview of central metabolic pathways for methane utilization in methylosinus trichosporium ob3b, a model alphaproteobacterial methanotro ...201323565111
distribution and origin of oxygen-dependent and oxygen-independent forms of mg-protoporphyrin monomethylester cyclase among phototrophic proteobacteria.magnesium-protoporphyrin ix monomethylester cyclase is one of the key enzymes of the bacteriochlorophyll biosynthesis pathway. there exist two fundamentally different forms of this enzyme. the oxygen-dependent form, encoded by the gene acsf, catalyzes the formation of the bacteriochlorophyll fifth ring using oxygen, whereas the oxygen-independent form encoded by the gene bche utilizes an oxygen atom extracted from water. the presence of acsf and bche genes was surveyed in various phototrophic pr ...201323396335
navigating the b(12) road: assimilation, delivery, and disorders of cobalamin.the reactivity of the cobalt-carbon bond in cobalamins is the key to their chemical versatility, supporting both methyl transfer and isomerization reactions. during evolution of higher eukaryotes that utilize vitamin b12, the high reactivity of the cofactor coupled with its low abundance pressured development of an efficient system for uptake, assimilation, and delivery of the cofactor to client b12-dependent enzymes. although most proteins suspected to be involved in b12 trafficking were discov ...201323539619
synchronous waves of failed soft sweeps in the laboratory: remarkably rampant clonal interference of alleles at a single locus.it has increasingly been recognized that adapting populations of microbes contain not one, but many lineages continually arising and competing at once. this process, termed "clonal interference," alters the rate and dynamics of adaptation and biases winning mutations toward those with the largest selective effect. here we uncovered a dramatic example of clonal interference between multiple similar mutations occurring at the same locus within replicate populations of methylobacterium extorquens a ...201323307898
isolation and expression of a cdna encoding methylmalonic aciduria type a protein from euglena gracilis z.in animals, cobalamin (cbl) is a cofactor for methionine synthase and methylmalonyl-coa mutase (mcm), which utilizes methylcobalamin and 5'-deoxyadenosylcobalamin (adocbl), respectively. the cbla complementation class of inborn errors of cbl metabolism in humans is one of three known disorders that affect adocbl synthesis. the gene responsible for cbla has been identified in humans (mmaa) as well as its homolog (meab) in methylobacterium extorquens. recently, it has been reported that human mmaa ...201324957894
fim-1, a new acquired metallo-β-lactamase from a pseudomonas aeruginosa clinical isolate from italy.acquired metallo-β-lactamases (mbls) are resistance determinants of increasing clinical importance in gram-negative bacterial pathogens, which confer a broad-spectrum β-lactam resistance, including carbapenems. several such enzymes have been described since the 1990s. in the present study, a novel acquired mbl, named fim-1, was identified and characterized. the bla(fim-1) gene was cloned from a multidrug-resistant pseudomonas aeruginosa clinical isolate (fi-14/157) cultured from a patient with a ...201323114762
draft genome sequence of methylobacterium sp. strain l2-4, a leaf-associated endophytic n-fixing bacterium isolated from jatropha curcas l.methylobacterium sp. strain l2-4 is an efficient nitrogen-fixing leaf colonizer of biofuel crop jatropha curcas. this strain is able to greatly improve the growth and seed yield of jatropha curcas and is the second reported genome sequence of plant growth-promoting bacteria isolated from jatropha curcas.201425502683
ethylmalonyl coenzyme a mutase operates as a metabolic control point in methylobacterium extorquens am1.the metabolism of one- and two-carbon compounds by the methylotrophic bacterium methylobacterium extorquens am1 involves high carbon flux through the ethylmalonyl coenzyme a (ethylmalonyl-coa) pathway (emc pathway). during growth on ethylamine, the emc pathway operates as a linear pathway carrying the full assimilatory flux to produce glyoxylate, malate, and succinate. assimilatory carbon enters the ethylmalonyl-coa pathway directly as acetyl-coa, bypassing pathways for formaldehyde oxidation/as ...201425448820
ethylmalonyl coenzyme a mutase operates as a metabolic control point in methylobacterium extorquens am1.the metabolism of one- and two-carbon compounds by the methylotrophic bacterium methylobacterium extorquens am1 involves high carbon flux through the ethylmalonyl coenzyme a (ethylmalonyl-coa) pathway (emc pathway). during growth on ethylamine, the emc pathway operates as a linear pathway carrying the full assimilatory flux to produce glyoxylate, malate, and succinate. assimilatory carbon enters the ethylmalonyl-coa pathway directly as acetyl-coa, bypassing pathways for formaldehyde oxidation/as ...201425448820
methylamine utilization via the n-methylglutamate pathway in methylobacterium extorquens pa1 involves a novel flow of carbon through c1 assimilation and dissimilation pathways.methylotrophs grow on reduced single-carbon compounds like methylamine as the sole source of carbon and energy. in methylobacterium extorquens am1, the best-studied aerobic methylotroph, a periplasmic methylamine dehydrogenase that catalyzes the primary oxidation of methylamine to formaldehyde has been examined in great detail. however, recent metagenomic data from natural ecosystems are revealing the abundance and importance of lesser-known routes, such as the n-methylglutamate pathway, for met ...201425225269
effective use of a horizontally-transferred pathway for dichloromethane catabolism requires post-transfer refinement.when microbes acquire new abilities through horizontal gene transfer, the genes and pathways must function under conditions with which they did not coevolve. if newly-acquired genes burden the host, their utility will depend on further evolutionary refinement of the recombinant strain. we used laboratory evolution to recapitulate this process of transfer and refinement, demonstrating that effective use of an introduced dichloromethane degradation pathway required one of several mutations to the ...201425418043
structure of the methanofuran/methanopterin-biosynthetic enzyme mj1099 from methanocaldococcus jannaschii.prior studies have indicated that mj1099 from methanocaldococcus jannaschii has roles in the biosynthesis of tetrahydromethanopterin and methanofuran, two key cofactors of one-carbon (c1) metabolism in diverse organisms including the methanogenic archaea. here, the structure of mj1099 has been solved to 1.7 å resolution using anomalous scattering methods. the results indicate that mj1099 is a member of the tim-barrel superfamily and that it is a homohexamer. bioinformatic analyses identified a p ...201425372812
probing the diversity of chloromethane-degrading bacteria by comparative genomics and isotopic fractionation.chloromethane (ch3cl) is produced on earth by a variety of abiotic and biological processes. it is the most important halogenated trace gas in the atmosphere, where it contributes to ozone destruction. current estimates of the global ch3cl budget are uncertain and suggest that microorganisms might play a more important role in degrading atmospheric ch3cl than previously thought. its degradation by bacteria has been demonstrated in marine, terrestrial, and phyllospheric environments. improving ou ...201425360131
contribution of silent mutations to thermal adaptation of rna bacteriophage qβ.changes in protein function and other biological properties, such as rna structure, are crucial for adaptation of organisms to novel or inhibitory environments. to investigate how mutations that do not alter amino acid sequence may be positively selected, we performed a thermal adaptation experiment using the single-stranded rna bacteriophage qβ in which the culture temperature was increased from 37.2°c to 41.2°c and finally to an inhibitory temperature of 43.6°c in a stepwise manner in three in ...201425056887
genetic and phenotypic comparison of facultative methylotrophy between methylobacterium extorquens strains pa1 and am1.methylobacterium extorquens am1, a strain serendipitously isolated half a century ago, has become the best-characterized model system for the study of aerobic methylotrophy (the ability to grow on reduced single-carbon compounds). however, with 5 replicons and 174 insertion sequence (is) elements in the genome as well as a long history of domestication in the laboratory, genetic and genomic analysis of m. extorquens am1 face several challenges. on the contrary, a recently isolated strain - m. ex ...201425232997
genome information of methylobacterium oryzae, a plant-probiotic methylotroph in the phyllosphere.pink-pigmented facultative methylotrophs in the rhizobiales are widespread in the environment, and many methylobacterium species associated with plants produce plant growth-promoting substances. to gain insights into the life style at the phyllosphere and the genetic bases of plant growth promotion, we determined and analyzed the complete genome sequence of methylobacterium oryzae cbmb20t, a strain isolated from rice stem. the genome consists of a 6.29-mb chromosome and four plasmids, designated ...201425211235
identification of a unique radical s-adenosylmethionine methylase likely involved in methanopterin biosynthesis in methanocaldococcus jannaschii.methanopterin (mpt) and its analogs are coenzymes required for methanogenesis and methylotrophy in specialized microorganisms. the methyl groups at c-7 and c-9 of the pterin ring distinguish mpt from all other pterin-containing natural products. however, the enzyme(s) responsible for the addition of these methyl groups has yet to be identified. here we demonstrate that a putative radical s-adenosyl-l-methionine (sam) enzyme superfamily member encoded by the mj0619 gene in the methanogen methanoc ...201425002541
ribosylhopane, a novel bacterial hopanoid, as precursor of c35 bacteriohopanepolyols in streptomyces coelicolor a3(2).wild-type streptomyces coelicolor a3(2) produces aminobacteriohopanetriol as the only elongated c35 hopanoid. the hopanoid phenotype of two mutants bearing a deletion of genes from a previously identified hopanoid biosynthesis gene cluster provides clues to the formation of c35 bacteriohopanepolyols. orf14 encodes a putative nucleosidase; its deletion induces the accumulation of adenosylhopane as it cannot be converted into ribosylhopane. orf18 encodes a putative transaminase; its deletion resul ...201425155017
repair rather than segregation of damage is the optimal unicellular aging strategy.how aging, being unfavourable for the individual, can evolve is one of the fundamental problems of biology. evidence for aging in unicellular organisms is far from conclusive. some studies found aging even in symmetrically dividing unicellular species; others did not find aging in the same, or in different, unicellular species, or only under stress. mathematical models suggested that segregation of non-genetic damage, as an aging strategy, would increase fitness. however, these models failed to ...201425184818
variation in the fitness effects of mutations with population density and size in escherichia coli.the fitness effects of mutations are context specific and depend on both external (e.g., environment) and internal (e.g., cellular stress, genetic background) factors. the influence of population size and density on fitness effects are unknown, despite the central role population size plays in the supply and fixation of mutations. we addressed this issue by comparing the fitness of 92 keio strains (escherichia coli k12 single gene knockouts) at comparatively high (1.2×10(7) cfus/ml) and low (2.5 ...201425121498
tunable control of polyproline helix (ppii) structure via aromatic electronic effects: an electronic switch of polyproline helix.aromatic rings exhibit defined interactions via the unique aromatic π face. aromatic amino acids interact favorably with proline residues via both the hydrophobic effect and aromatic-proline interactions, c-h/π interactions between the aromatic π face and proline ring c-h bonds. the canonical aromatic amino acids trp, tyr, and phe strongly disfavor a polyproline helix (ppii) when they are present in proline-rich sequences because of the large populations of cis amide bonds induced by favorable a ...201425075447
evolution of escherichia coli to 42 °c and subsequent genetic engineering reveals adaptive mechanisms and novel mutations.adaptive laboratory evolution (ale) has emerged as a valuable method by which to investigate microbial adaptation to a desired environment. here, we performed ale to 42 °c of ten parallel populations of escherichia coli k-12 mg1655 grown in glucose minimal media. tightly controlled experimental conditions allowed selection based on exponential-phase growth rate, yielding strains that uniformly converged toward a similar phenotype along distinct genetic paths. adapted strains possessed as few as ...201425015645
emergent biosynthetic capacity in simple microbial communities.microbes have an astonishing capacity to transform their environments. yet, the metabolic capacity of a single species is limited and the vast majority of microorganisms form complex communities and join forces to exhibit capabilities far exceeding those achieved by any single species. such enhanced metabolic capacities represent a promising route to many medical, environmental, and industrial applications and call for the development of a predictive, systems-level understanding of synergistic m ...201424992662
phylogeny poorly predicts the utility of a challenging horizontally transferred gene in methylobacterium strains.horizontal gene transfer plays a crucial role in microbial evolution. while much is known about the mechanisms that determine whether physical dna can be transferred into a new host, the factors determining the utility of the transferred genes are less clear. we have explored this issue using dichloromethane consumption in methylobacterium strains. methylobacterium extorquens dm4 expresses a dichloromethane dehalogenase (dcma) that has been acquired through horizontal gene transfer and allows th ...201424682326
trade-offs drive resource specialization and the gradual establishment of ecotypes.speciation is driven by many different factors. among those are trade-offs between different ways an organism utilizes resources, and these trade-offs can constrain the manner in which selection can optimize traits. limited migration among allopatric populations and species interactions can also drive speciation, but here we ask if trade-offs alone are sufficient to drive speciation in the absence of other factors.201424885598
metabolomic strategies for the identification of new enzyme functions and metabolic pathways.recent technological advances in accurate mass spectrometry and data analysis have revolutionized metabolomics experimentation. activity-based and global metabolomic profiling methods allow simultaneous and rapid screening of hundreds of metabolites from a variety of chemical classes, making them useful tools for the discovery of novel enzymatic activities and metabolic pathways. by using the metabolome of the relevant organism or close species, these methods capitalize on biological relevance, ...201424829223
metabolic resource allocation in individual microbes determines ecosystem interactions and spatial dynamics.the interspecies exchange of metabolites plays a key role in the spatiotemporal dynamics of microbial communities. this raises the question of whether ecosystem-level behavior of structured communities can be predicted using genome-scale metabolic models for multiple organisms. we developed a modeling framework that integrates dynamic flux balance analysis with diffusion on a lattice and applied it to engineered communities. first, we predicted and experimentally confirmed the species ratio to w ...201424794435
genomic and transcriptomic analyses of the facultative methanotroph methylocystis sp. strain sb2 grown on methane or ethanol.a minority of methanotrophs are able to utilize multicarbon compounds as growth substrates in addition to methane. the pathways utilized by these microorganisms for assimilation of multicarbon compounds, however, have not been explicitly examined. here, we report the draft genome of the facultative methanotroph methylocystis sp. strain sb2 and perform a detailed transcriptomic analysis of cultures grown with either methane or ethanol. evidence for use of the canonical methane oxidation pathway a ...201424610846
methylobacterium and its role in health care-associated infection.methylobacterium species are a cause of health care-associated infection, including infections in immunocompromised hosts. the ability of methylobacterium species to form biofilms and to develop resistance to high temperatures, drying, and disinfecting agents may explain the colonization of methylobacterium in the hospital environment in, e.g., endoscopes. due to its slow growth, it can be easily missed during microbiological surveillance of endoscope reprocessing. the purpose of this minireview ...201424430456
new vectors for chromosomal integration enable high-level constitutive or inducible magnetosome expression of fusion proteins in magnetospirillum gryphiswaldense.the alphaproteobacterium magnetospirillum gryphiswaldense biomineralizes magnetosomes, which consist of monocrystalline magnetite cores enveloped by a phospholipid bilayer containing specific proteins. magnetosomes represent magnetic nanoparticles with unprecedented magnetic and physicochemical characteristics. these make them potentially useful in a number of biotechnological and biomedical applications. further functionalization can be achieved by expression of foreign proteins via genetic fus ...201424532068
application of stable isotope-assisted metabolomics for cell metabolism studies.the applications of stable isotopes in metabolomics have facilitated the study of cell metabolisms. stable isotope-assisted metabolomics requires: (1) properly designed tracer experiments; (2) stringent sampling and quenching protocols to minimize isotopic alternations; (3) efficient metabolite separations; (4) high resolution mass spectrometry to resolve overlapping peaks and background noises; and (5) data analysis methods and databases to decipher isotopic clusters over a broad m/z range (mas ...201424957020
mapping the fitness landscape of gene expression uncovers the cause of antagonism and sign epistasis between adaptive mutations.how do adapting populations navigate the tensions between the costs of gene expression and the benefits of gene products to optimize the levels of many genes at once? here we combined independently-arising beneficial mutations that altered enzyme levels in the central metabolism of methylobacterium extorquens to uncover the fitness landscape defined by gene expression levels. we found strong antagonism and sign epistasis between these beneficial mutations. mutations with the largest individual b ...201424586190
stable isotope-labeling studies in metabolomics: new insights into structure and dynamics of metabolic networks.the rapid emergence of metabolomics has enabled system-wide measurements of metabolites in various organisms. however, advances in the mechanistic understanding of metabolic networks remain limited, as most metabolomics studies cannot routinely provide accurate metabolite identification, absolute quantification and flux measurement. stable isotope labeling offers opportunities to overcome these limitations. here we describe some current approaches to stable isotope-labeled metabolomics and provi ...201424568354
laboratory divergence of methylobacterium extorquens am1 through unintended domestication and past selection for antibiotic resistance.a common assumption of microorganisms is that laboratory stocks will remain genetically and phenotypically constant over time, and across laboratories. it is becoming increasingly clear, however, that mutations can ruin strain integrity and drive the divergence or "domestication" of stocks. since its discovery in 1960, a stock of methylobacterium extorquens am1 ("am1") has remained in the lab, propagated across numerous growth and storage conditions, researchers, and facilities. to explore the e ...201424384040
pentanol and benzyl alcohol attack bacterial surface structures differently.the genus methylobacterium tolerates hygiene agents like benzalkonium chloride (bac), and infection with this organism is an important public health issue. here, we found that the combination of bac with particular alcohols at nonlethal concentrations in terms of their solitary uses significantly reduced bacterial viability after only 5 min of exposure. among the alcohols, raman spectroscopic analyses showed that pentanol (pentyl alcohol [pea]) and benzyl alcohol (bza) accelerated the cellular a ...201526519389
pentanol and benzyl alcohol attack bacterial surface structures differently.the genus methylobacterium tolerates hygiene agents like benzalkonium chloride (bac), and infection with this organism is an important public health issue. here, we found that the combination of bac with particular alcohols at nonlethal concentrations in terms of their solitary uses significantly reduced bacterial viability after only 5 min of exposure. among the alcohols, raman spectroscopic analyses showed that pentanol (pentyl alcohol [pea]) and benzyl alcohol (bza) accelerated the cellular a ...201526519389
isolation of radiation-resistant bacteria from mars analog antarctic dry valleys by preselection, and the correlation between radiation and desiccation resistance.extreme radiation-resistant microorganisms can survive doses of ionizing radiation far greater than are present in the natural environment. radiation resistance is believed to be an incidental adaptation to desiccation resistance, as both hazards cause similar cellular damage. desert soils are, therefore, promising targets to prospect for new radiation-resistant strains. this is the first study to isolate radiation-resistant microbes by using gamma-ray exposure preselection from the extreme cold ...201526684506
identification of two mutations increasing the methanol tolerance of corynebacterium glutamicum.methanol is present in most ecosystems and may also occur in industrial applications, e.g. as an impurity of carbon sources such as technical glycerol. methanol often inhibits growth of bacteria, thus, methanol tolerance may limit fermentative production processes.201526474849
roles of the crotonyl-coa carboxylase/reductase homologues in acetate assimilation and biosynthesis of immunosuppressant fk506 in streptomyces tsukubaensis.in microorganisms lacking a functional glyoxylate cycle, acetate can be assimilated by alternative pathways of carbon metabolism such as the ethylmalonyl-coa (emc) pathway. among the enzymes converting coa-esters of the emc pathway, there is a unique carboxylase that reductively carboxylates crotonyl-coa, crotonyl-coa carboxylase/reductase (ccr). in addition to the emc pathway, gene homologues of ccr can be found in secondary metabolite gene clusters that are involved in the provision of structu ...201526466669
cerium regulates expression of alternative methanol dehydrogenases in methylosinus trichosporium ob3b.methanotrophs have multiple methane monooxygenases that are well known to be regulated by copper, i.e., a "copper switch." at low copper/biomass ratios the soluble methane monooxygenase (smmo) is expressed while expression and activity of the particulate methane monooxygenase (pmmo) increases with increasing availability of copper. in many methanotrophs there are also multiple methanol dehydrogenases (medhs), one based on mxa and another based on xox. mxa-medh is known to have calcium in its act ...201526296730
diversity, bacterial symbionts and antibacterial potential of gut-associated fungi isolated from the pantala flavescens larvae in china.the diversity of fungi associated with the gut of pantala flavescens larvae was investigated using a culture-dependent method and molecular identification based on an analysis of the internally transcribed spacer sequence. in total, 48 fungal isolates were obtained from p. flavescens larvae. based on phylogenetic analyses, the fungal isolates were grouped in 5 classes and 12 different genera. fourteen bacterial 16s rdna sequences derived from total genomic dna extractions of fungal mycelia were ...201526221957
proteins related to the type i secretion system are associated with secondary seca_dead domain proteins in some species of planctomycetes, verrucomicrobia, proteobacteria, nitrospirae and chlorobi.a number of bacteria belonging to the pvc (planctomycetes-verrucomicrobia-chlamydiae) super-phylum contain unusual ribosome-bearing intracellular membranes. the evolutionary origins and functions of these membranes are unknown. some proteins putatively associated with the presence of intracellular membranes in pvc bacteria contain signal peptides. signal peptides mark proteins for translocation across the cytoplasmic membrane in prokaryotes, and the membrane of the endoplasmic reticulum in eukar ...201526030905
pivotal roles of phyllosphere microorganisms at the interface between plant functioning and atmospheric trace gas dynamics.the phyllosphere, which lato sensu consists of the aerial parts of plants, and therefore primarily, of the set of photosynthetic leaves, is one of the most prevalent microbial habitats on earth. phyllosphere microbiota are related to original and specific processes at the interface between plants, microorganisms and the atmosphere. recent -omics studies have opened fascinating opportunities for characterizing the spatio-temporal structure of phyllosphere microbial communities in relation with st ...201526052316
methenyl-dephosphotetrahydromethanopterin is a regulatory signal for acclimation to changes in substrate availability in methylobacterium extorquens am1.during an environmental perturbation, the survival of a cell and its response to the perturbation depend on both the robustness and functionality of the metabolic network. the regulatory mechanisms that allow the facultative methylotrophic bacterium methylobacterium extorquens am1 to effect the metabolic transition from succinate to methanol growth are not well understood. methenyl-dephosphotetrahydromethanopterin (methenyl-dh4mpt), an early intermediate during methanol metabolism, transiently a ...201525845846
interactions of methylotrophs with plants and other heterotrophic bacteria.methylotrophs, which can utilize methane and/or methanol as sole carbon and energy sources, are key players in the carbon cycle between methane and co₂, the two most important greenhouse gases. this review describes the relationships between methylotrophs and plants, and between methanotrophs (methane-utilizers, a subset of methylotrophs) and heterotrophic bacteria. some plants emit methane and methanol from their leaves, and provide methylotrophs with habitats. methanol-utilizing methylotrophs ...201527682083
crystal structures of mycobacterial meab and mmaa-like gtpases.the methylmalonyl co-a mutase-associated gtpase meab from methylobacterium extorquens is involved in glyoxylate regulation and required for growth. in humans, mutations in the homolog methylmalonic aciduria associated protein (mmaa) cause methylmalonic aciduria, which is often fatal. the central role of meab from bacteria to humans suggests that meab is also important in other, pathogenic bacteria such as mycobacterium tuberculosis. however, the identity of the mycobacterial meab homolog is pres ...201525832174
sulfur isotope fractionation during the evolutionary adaptation of a sulfate-reducing bacterium.dissimilatory sulfate reduction is a microbial catabolic pathway that preferentially processes less massive sulfur isotopes relative to their heavier counterparts. this sulfur isotope fractionation is recorded in ancient sedimentary rocks and generally is considered to reflect a phenotypic response to environmental variations rather than to evolutionary adaptation. modern sulfate-reducing microorganisms isolated from similar environments can exhibit a wide range of sulfur isotope fractionations, ...201525662968
experimental horizontal gene transfer of methylamine dehydrogenase mimics prevalent exchange in nature and overcomes the methylamine growth constraints posed by the sub-optimal n-methylglutamate pathway.methylamine plays an important role in the global carbon and nitrogen budget; microorganisms that grow on reduced single carbon compounds, methylotrophs, serve as a major biological sink for methylamine in aerobic environments. two non-orthologous, functionally degenerate routes for methylamine oxidation have been studied in methylotrophic proteobacteria: methylamine dehydrogenase and the n-methylglutamate pathway. recent work suggests the n-methylglutamate (nmg) pathway may be more common in na ...201527682079
transhydrogenase promotes the robustness and evolvability of e. coli deficient in nadph production.metabolic networks revolve around few metabolites recognized by diverse enzymes and involved in myriad reactions. though hub metabolites are considered as stepping stones to facilitate the evolutionary expansion of biochemical pathways, changes in their production or consumption often impair cellular physiology through their system-wide connections. how does metabolism endure perturbations brought immediately by pathway modification and restore hub homeostasis in the long run? to address this qu ...201525715029
role of extracytoplasmic function sigma factors in biofilm formation of porphyromonas gingivalis.porphyromonas gingivalis has been implicated as a major pathogen in the development and progression of chronic periodontitis. p. gingivalis biofilm formation in the subgingival crevice plays an important role in the ability of the bacteria to tolerate stress signals outside the cytoplasmic membrane. some bacteria use a distinct subfamily of sigma factors to regulate their extracytoplasmic functions (the ecf subfamily). the objective of this study was to determine if p. gingivalis ecf sigma facto ...201525596817
a mutational hotspot and strong selection contribute to the order of mutations selected for during escherichia coli adaptation to the gut.the relative role of drift versus selection underlying the evolution of bacterial species within the gut microbiota remains poorly understood. the large sizes of bacterial populations in this environment suggest that even adaptive mutations with weak effects, thought to be the most frequently occurring, could substantially contribute to a rapid pace of evolutionary change in the gut. we followed the emergence of intra-species diversity in a commensal escherichia coli strain that previously acqui ...201627812114
love the one you're with: replicate viral adaptations converge on the same phenotypic change.parallelism is important because it reveals how inherently stochastic adaptation is. even as we come to better understand evolutionary forces, stochasticity limits how well we can predict evolutionary outcomes. here we sought to quantify parallelism and some of its underlying causes by adapting a bacteriophage (id11) with nine different first-step mutations, each with eight-fold replication, for 100 passages. this was followed by whole-genome sequencing five isolates from each endpoint. a large ...201627547540
difference in c3-c4 metabolism underlies tradeoff between growth rate and biomass yield in methylobacterium extorquens am1.two variants of methylobacterium extorquens am1 demonstrated a trade-off between growth rate and biomass yield. in addition, growth rate and biomass yield were also affected by supplementation of growth medium with different amounts of cobalt. the metabolism changes relating to these growth phenomena as well as the trade-off were investigated in this study. (13)c metabolic flux analysis was used to generate a detailed central carbon metabolic flux map with both absolute and normalized flux value ...201627435978
transfer of a catabolic pathway for chloromethane in methylobacterium strains highlights different limitations for growth with chloromethane or with dichloromethane.chloromethane (cm) is an ozone-depleting gas, produced predominantly from natural sources, that provides an important carbon source for microbes capable of consuming it. cm catabolism has been difficult to study owing to the challenging genetics of its native microbial hosts. since the pathways for cm catabolism show evidence of horizontal gene transfer, we reproduced this transfer process in the laboratory to generate new cm-catabolizing strains in tractable hosts. we demonstrate that six putat ...201627486448
the frequency of internal shine-dalgarno-like motifs in prokaryotes.in prokaryotes, translation initiation typically depends on complementary binding between a g-rich shine-dalgarno (sd) motif in the 5' untranslated region of mrnas, and the 3' tail of the 16s ribosomal rna (the anti-sd sequence). in some cases, internal sd-like motifs in the coding region generate "programmed" ribosomal pauses that are beneficial for protein folding or accurate targeting. on the other hand, such pauses can also reduce protein production, generating purifying selection against in ...201627189998
metabolomics revealed an association of metabolite changes and defective growth in methylobacterium extorquens am1 overexpressing ecm during growth on methanol.methylobacterium extorquens am1 is a facultative methylotroph capable of growth on both single-carbon and multi-carbon compounds. the ethylmalonyl-coa (emc) pathway is one of the central assimilatory pathways in m. extorquens during growth on c1 and c2 substrates. previous studies had shown that ethylmalonyl-coa mutase functioned as a control point during the transition from growth on succinate to growth on ethylamine. in this study we overexpressed ecm, phaa, mcmab and found that upregulating e ...201627116459
shifts in plant foliar and floral metabolomes in response to the suppression of the associated microbiota.the phyllospheric microbiota is assumed to play a key role in the metabolism of host plants. its role in determining the epiphytic and internal plant metabolome, however, remains to be investigated. we analyzed the liquid chromatography-mass spectrometry (lc-ms) profiles of the epiphytic and internal metabolomes of the leaves and flowers of sambucus nigra with and without external antibiotic treatment application.201627048394
epistasis and the structure of fitness landscapes: are experimental fitness landscapes compatible with fisher's geometric model?the fitness landscape defines the relationship between genotypes and fitness in a given environment and underlies fundamental quantities such as the distribution of selection coefficient and the magnitude and type of epistasis. a better understanding of variation in landscape structure across species and environments is thus necessary to understand and predict how populations will adapt. an increasing number of experiments investigate the properties of fitness landscapes by identifying mutations ...201627052568
xoxf acts as the predominant methanol dehydrogenase in the type i methanotroph methylomicrobium buryatense.many methylotrophic taxa harbor two distinct methanol dehydrogenase (mdh) systems for oxidizing methanol to formaldehyde: the well-studied calcium-dependent mxafi type and the more recently discovered lanthanide-containing xoxf type. mxafi has traditionally been accepted as the major functional mdh in bacteria that contain both enzymes. however, in this study, we present evidence that, in a type i methanotroph, methylomicrobium buryatense, xoxf is likely the primary functional mdh in the environ ...201626858104
lanthanide-dependent regulation of methanol oxidation systems in methylobacterium extorquens am1 and their contribution to methanol growth.methylobacterium extorquens am1 has two distinct types of methanol dehydrogenase (medh) enzymes that catalyze the oxidation of methanol to formaldehyde. mxafi-medh requires pyrroloquinoline quinone (pqq) and ca in its active site, while xoxf-medh requires pqq and lanthanides, such as ce and la. using medh mutant strains to conduct growth analysis and medh activity assays, we demonstrate that m. extorquens am1 has at least one additional lanthanide-dependent methanol oxidation system contributing ...201626833413
associations between ectomycorrhizal fungi and bacterial needle endophytes in pinus radiata: implications for biotic selection of microbial communities.studies of the ecological and evolutionary relationships between plants and their associated microbes have long been focused on single microbes, or single microbial guilds, but in reality, plants associate with a diverse array of microbes from a varied set of guilds. as such, multitrophic interactions among plant-associated microbes from multiple guilds represent an area of developing research, and can reveal how complex microbial communities are structured around plants. interactions between co ...201627065966
comprehensive genomic analyses of the om43 clade, including a novel species from the red sea, indicate ecotype differentiation among marine methylotrophs.the om43 clade within the family methylophilaceae of betaproteobacteria represents a group of methylotrophs that play important roles in the metabolism of c1 compounds in marine environments and other aquatic environments around the globe. using dilution-to-extinction cultivation techniques, we successfully isolated a novel species of this clade (here designated mbrs-h7) from the ultraoligotrophic open ocean waters of the central red sea. phylogenomic analyses indicate that mbrs-h7 is a novel sp ...201626655752
biosynthesis of polyhydroxyalkanoate copolymers from methanol by methylobacterium extorquens am1 and the engineered strains under cobalt-deficient conditions.methylobacterium extorquens am1 has been shown to accumulate polyhydroxyalkanoate (pha) composed solely of (r)-3-hydroxybutyrate (3hb) during methylotrophic growth. the present study demonstrated that the wild-type strain am1 grown under co²⁺-deficient conditions accumulated copolyesters of 3hb and a c₅-monomer, (r)-3-hydroxyvalerate (3hv), using methanol as the sole carbon source. the 3hv unit was supposed to be derived from propionyl-coa, synthesized via the ethylmalonyl-coa pathway impaired b ...201424430207
thioesterases for ethylmalonyl-coa pathway derived dicarboxylic acid production in methylobacterium extorquens am1.the ethylmalonyl-coenzyme a pathway (emcp) is a recently discovered pathway present in diverse α-proteobacteria such as the well studied methylotroph methylobacterium extorquens am1. its glyoxylate regeneration function is obligatory during growth on c1 carbon sources like methanol. the emcp contains special coa esters, of which dicarboxylic acid derivatives are of high interest as building blocks for chemical industry. the possible production of dicarboxylic acids out of the alternative, non-fo ...201424419796
bioconversion of methanol to value-added mevalonate by engineered methylobacterium extorquens am1 containing an optimized mevalonate pathway.methylotrophic biosynthesis using methanol as a feedstock is a promising and attractive method to solve the over-dependence of the bioindustry on sugar feedstocks derived from grains that are used for food. in this study, we introduced and engineered the mevalonate pathway into methylobacterium extorquens am1 to achieve high mevalonate production from methanol, which could be a platform for terpenoid synthesis. we first constructed a natural operon (mve) harboring the mvas and mvae genes from en ...201626521242
(1)h, (13)c, and (15)n resonance assignments and secondary structure information for methylobacterium extorquens pqqd and the complex of pqqd with pqqa.the ribosomally synthesized and post-translationally modified peptide (ripp), pyrroloquinoline quinone (pqq), is a dehydrogenase cofactor synthesized by, but not exclusively used by, certain prokaryotes. ripps represent a rapidly expanding and diverse class of natural products-many of which have therapeutic potential-and the biosynthetic pathways for these are gaining attention. five gene products from the pqq operon (pqqa, pqqb, pqqc, pqqd, and pqqe) are essential for pqq biosynthesis. the subs ...201627638737
adding biotic complexity alters the metabolic benefits of mutualism.mutualism is ubiquitous in nature and plays an integral role in most communities. to predict the eco-evolutionary dynamics of mutualism it is critical to extend classic pair-wise analysis to include additional species. we investigated the effect of adding a third species to a pair-wise mutualism in a spatially structured environment. we tested the hypotheses that selection for costly excretions in a focal population (i) decreases when an exploiter is added (ii) increases when a third mutualist i ...201627272242
hopanoid-free methylobacterium extorquens dm4 overproduces carotenoids and has widespread growth impairment.hopanoids are sterol-like membrane lipids widely used as geochemical proxies for bacteria. currently, the physiological role of hopanoids is not well understood, and this represents one of the major limitations in interpreting the significance of their presence in ancient or contemporary sediments. previous analyses of mutants lacking hopanoids in a range of bacteria have revealed a range of phenotypes under normal growth conditions, but with most having at least an increased sensitivity to toxi ...201728319163
structure of methylobacterium extorquens malyl-coa lyase: coa-substrate binding correlates with domain shift.malyl-coa lyase (mcl) is an mg(2+)-dependent enzyme that catalyzes the reversible cleavage of (2s)-4-malyl-coa to yield acetyl-coa and glyoxylate. mcl enzymes, which are found in a variety of bacteria, are members of the citrate lyase-like family and are involved in the assimilation of one- and two-carbon compounds. here, the 1.56 å resolution x-ray crystal structure of mcl from methylobacterium extorquens am1 with bound mg(2+) is presented. structural alignment with the closely related rhodobac ...201728177317
regulation of root development in arabidopsis thaliana by phytohormone-secreting epiphytic methylobacteria.in numerous experimental studies, seedlings of the model dicot arabidopsis thaliana have been raised on sterile mineral salt agar. however, under natural conditions, no plant has ever grown in an environment without bacteria. here, we document that germ-free (gnotobiotic) seedlings, raised on mineral salt agar without sucrose, develop very short root hairs. in the presence of a soil extract that contains naturally occurring microbes, root hair elongation is promoted; this effect can be mimicked ...201728054231
biosensor-assisted transcriptional regulator engineering for methylobacterium extorquens am1 to improve mevalonate synthesis by increasing the acetyl-coa supply.acetyl-coa is not only an important intermediate metabolite for cells but also a significant precursor for production of industrially interesting metabolites. methylobacterium extorquens am1, a model strain of methylotrophic cell factories using methanol as carbon source, is of interest because it produces abundant coenzyme a compounds capable of directing to synthesis of different useful compounds from methanol. however, acetyl-coa is not always efficiently accumulated in m. extorquens am1, as ...201727919791
production of 2-hydroxyisobutyric acid from methanol by methylobacterium extorquens am1 expressing (r)-3-hydroxybutyryl coenzyme a-isomerizing enzymes.the biotechnological production of the methyl methacrylate precursor 2-hydroxyisobutyric acid (2-hiba) via bacterial poly-3-hydroxybutyrate (phb) overflow metabolism requires suitable (r)-3-hydroxybutyryl coenzyme a (coa)-specific coenzyme b12-dependent mutases (rcm). here, we characterized a predicted mutase from bacillus massiliosenegalensis jc6 as a mesophilic rcm closely related to the thermophilic enzyme previously identified in kyrpidia tusciae dsm 2912 (m.-t. weichler et al., appl environ ...201727836853
pyrroloquinoline quinone ethanol dehydrogenase in methylobacterium extorquens am1 extends lanthanide-dependent metabolism to multicarbon substrates.lanthanides are utilized by microbial methanol dehydrogenases, and it has been proposed that lanthanides may be important for other type i alcohol dehydrogenases. a triple mutant strain (mxaf xoxf1 xoxf2; named mdh-3), deficient in the three known methanol dehydrogenases of the model methylotroph methylobacterium extorquens am1, is able to grow poorly with methanol if exogenous lanthanides are added to the growth medium. when the gene encoding a putative quinoprotein ethanol dehydrogenase, exaf, ...201627573017
systems-level proteomics of two ubiquitous leaf commensals reveals complementary adaptive traits for phyllosphere colonization.plants are colonized by a diverse community of microorganisms, the plant microbiota, exhibiting a defined and conserved taxonomic structure. niche separation based on spatial segregation and complementary adaptation strategies likely forms the basis for coexistence of the various microorganisms in the plant environment. to gain insights into organism-specific adaptations on a molecular level, we selected two exemplary community members of the core leaf microbiota and profiled their proteomes upo ...201627457762
towards stable kinetics of large metabolic networks: nonequilibrium potential function approach.while the biochemistry of metabolism in many organisms is well studied, details of the metabolic dynamics are not fully explored yet. acquiring adequate in vivo kinetic parameters experimentally has always been an obstacle. unless the parameters of a vast number of enzyme-catalyzed reactions happened to fall into very special ranges, a kinetic model for a large metabolic network would fail to reach a steady state. in this work we show that a stable metabolic network can be systematically establi ...201627415300
the arabidopsis leaf transcriptome reveals distinct but also overlapping responses to colonization by phyllosphere commensals and pathogen infection with impact on plant health.plants are colonized by a variety of bacteria, most of which are not pathogenic. currently, the plant responses to phyllosphere commensals or to pathogen infection in the presence of commensals are not well understood. here, we examined the transcriptional response of arabidopsis thaliana leaves to colonization by common commensal bacteria in a gnotobiotic system using rna sequencing and conducted plant mutant assays. arabidopsis responded differently to the model bacteria sphingomonas melonis f ...201627306148
[enhanced resistance of pea plants to oxidative: stress caused by paraquat during colonization by aerobic methylobacteria].the influence of colonization of the pea (pisum sativum l.) by aerobic methylobacteria of five different species (methylophilus flavus ship, methylobacterium extorquens g10, methylobacillus arboreus iva, methylopila musalis musa, methylopila turkiensis sidel) on plant resistance to paraquat-induced stresses has been studied. the normal conditions of pea colonization by methylobacteria were characterized by a decrease in the activity of antioxidant enzymes (superoxide dismutase, catalase, and per ...201627266250
selection maintains apparently degenerate metabolic pathways due to tradeoffs in using methylamine for carbon versus nitrogen.microorganisms often encode multiple non-orthologous metabolic modules that catalyze the same reaction. however, little experimental evidence actually demonstrates a selective basis for metabolic degeneracy. many methylotrophs-microorganisms that grow on reduced single-carbon compounds-like methylobacterium extorquens am1 encode two routes for methylamine oxidation: the periplasmic methylamine dehydrogenase (madh) and the cytoplasmic n-methylglutamate (nmg) pathway. in methylobacterium extorquen ...201627212407
calorespirometric feeding control enhances bioproduction from toxic feedstocks-demonstration for biopolymer production out of methanol.the sustainable production of fuels and industrial bulk chemicals by microorganisms in biotechnological processes is promising but still facing various challenges. in particular, toxic substrates require an efficient process control strategy. methanol, as an example, has the potential to become a major future feedstock due to its availability from fossil and renewable resources. however, besides being toxic, methanol is highly volatile. to optimize its dosage during microbial cultivations, an in ...201627043974
[functionality of the xoxf gene in methylobacterium dichloromethanicum dm4].activation of expression of the xoxfgene encoding pqq-dependent methanol/ethanol dehydrogenase (metdi2492) in dichloromethane- (dcm) -grown methylobacterium dichloromethanicum dm4 was first demonstrated. the sequence of the only xoxf homolog found in the genome of strain dm4 exhibited 50% identity to that of the protein (mxaf) of the large subunit of methanol dehydrogenase (mdh). a knockout mutant with the inactivate xoxf gene (δxoxf) was found to be unable to grow on methanol due to the absence ...201626964359
the one-carbon carrier methylofuran from methylobacterium extorquens am1 contains a large number of α- and γ-linked glutamic acid residues.methylobacterium extorquens am1 uses dedicated cofactors for one-carbon unit conversion. based on the sequence identities of enzymes and activity determinations, a methanofuran analog was proposed to be involved in formaldehyde oxidation in alphaproteobacteria. here, we report the structure of the cofactor, which we termed methylofuran. using an in vitro enzyme assay and lc-ms, methylofuran was identified in cell extracts and further purified. from the exact mass and ms-ms fragmentation pattern, ...201626895963
non-targeted determination of (13)c-labeling in the methylobacterium extorquens am1 metabolome using the two-dimensional mass cluster method and principal component analysis.a novel analytical workflow is presented for the analysis of time-dependent (13)c-labeling of the metabolites in the methylotrophic bacterium methylobacterium extorquens am1 using gas chromatography time-of-flight mass spectrometry (gc-tofms). using (13)c-methanol as the substrate in a time course experiment, the method provides an accurate determination of the number of carbons converted to the stable isotope. the method also extracts a quantitative isotopic dilution time course profile for (13 ...201626787164
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