the paradoxical cyanide-stimulated respiration of zymomonas mobilis: cyanide sensitivity of alcohol dehydrogenase (adh ii).the respiratory inhibitor cyanide stimulates growth of the ethanologenic bacterium zymomonas mobilis, perhaps by diverting reducing equivalents from respiration to ethanol synthesis, thereby minimizing accumulation of toxic acetaldehyde. this study sought to identify cyanide-sensitive components of respiration. in aerobically grown, permeabilized z. mobilis cells, addition of 200 microm cyanide caused gradual inhibition of adh ii, the iron-containing alcohol dehydrogenase isoenzyme, which, in ae ...200312855725
expression of galp and glk in a escherichia coli pts mutant restores glucose transport and increases glycolytic flux to fermentation escherichia coli, the uptake and phosphorylation of glucose is carried out mainly by the phosphotransferase system (pts). despite the efficiency of glucose transport by pts, the required consumption of 1 mol of phosphoenolpyruvate (pep) for each mol of internalized glucose represents a drawback for some biotechnological applications where pep is a precursor of the desired product. for this reason, there is considerable interest in the generation of strains that can transport glucose efficient ...200312889033
screening of yeasts for cell-free production of (r)-phenylacetylcarbinol.105 yeast strains from 10 genera and 40 species were evaluated for cell-free production of (r)-phenylacetylcarbinol (pac), the chiral precursor in the manufacture of the pharmaceuticals ephedrine and pseudoephedrine. carboligase activity of pyruvate decarboxylase (pdc), forming pac from benzaldehyde and pyruvate, was found in extracts of 98 strains. pac was not formed from benzaldehyde and acetaldehyde, an activity of bacterial pdcs from zymomonas mobilis and zymobacter palmae. two interesting g ...200312889791
an essential role for aspartate 264 in catalysis by trna-guanine transglycosylase from escherichia coli.trna-guanine transglycosylase (tgt) catalyzes a post-transcriptional base-exchange reaction involved in the incorporation of the modified base queuine (q) into the wobble position of certain trnas. catalysis by tgt occurs through a double-displacement mechanism that involves the formation of a covalent enzyme-rna intermediate (kittendorf, j. d., barcomb, l. m., nonekowski, s. t., and garcia, g. a. (2001) biochemistry 40, 14123-14133). the tgt chemical mechanism requires the protonation of the di ...200312909636
synthesis of the aminocyclopentitol moieties of the hopanoids of zymomonas mobilis and 'anacystis montana'.the first synthesis of the cyclopentitol units in bacterial hopanoids has been accomplished from d-glucosamine and the possible biological activity of the free cyclitols as glycosidase inhibitors has been studied.200312932010
chemical trapping and crystal structure of a catalytic trna guanine transglycosylase covalent intermediate.prokaryotic trna guanine transglycosylase (tgt) catalyzes replacement of guanine (g) by 7-aminomethyl-7-deazaguanine (preq1) at the wobble position of four specific trnas. addition of 9-deazaguanine (9dzg) to a reaction mixture of zymomonas mobilis tgt and an rna substrate allowed us to trap, purify and crystallize a chemically competent covalent intermediate of the tgt-catalyzed reaction. the crystal structure of the tgt-rna-9dzg ternary complex at a resolution of 2.9 a reveals, unexpectedly, t ...200312949492
a study of the cider-sickness bacillus; a new variety of zymomonas anaerobia. 195613385435
bacteria engineered for fuel ethanol production: current status.the lack of industrially suitable microorganisms for converting biomass into fuel ethanol has traditionally been cited as a major technical roadblock to developing a bioethanol industry. in the last two decades, numerous microorganisms have been engineered to selectively produce ethanol. lignocellulosic biomass contains complex carbohydrates that necessitate utilizing microorganisms capable of fermenting sugars not fermentable by brewers' yeast. the most significant of these is xylose. the great ...200313680206
comparative catabolism of carbohydrates in pseudomonas species. 196013834438
influence of aeration and of pantothenate on growth yields of zymomonas mobilis.belaïch, jean-pierre (centre national de la recherche scientifique, marseille, france), and jacques c. senez. influence of aeration and of pantothenate on growth yields of zymomonas mobilis. j. bacteriol. 89:1195-1200. 1965.-the growth yields and rates of zymomonas mobilis were measured in aerobic and anaerobic cultures on glucose medium containing yeast extract, amino acids, or ammonium chloride as the nitrogen source. in the absence of yeast extract, pantothenate was required. the growth yield ...196514292985
glycolytic enzymes in zymomonas mobilis.raps, shirley (university of illinois, urbana) and r. d. demoss. glycolytic enzymes in zymomonas mobilis. j. bacteriol. 84:115-118. 1962-an enzyme extract of zymomonas mobilis (pseudomonas lindneri) was capable of fermenting glucose-6-phosphate to co(2) and ethanol. the extract was found to contain phosphohexoisomerase, aldolase, and glyceraldehyde-3-phosphate dehydrogenase, but no demonstrable phosphohexokinase. the lack of isotope-mixing found in earlier studies is, thus, explained on an enzym ...196214490400
flexible adaptations in the structure of the trna-modifying enzyme trna-guanine transglycosylase and their implications for substrate selectivity, reaction mechanism and structure-based drug design.the enzyme trna-guanine transglycosylase (tgt, ec catalyses a base-exchange reaction that leads to anticodon modifications of certain trnas. the tgt enzymes of the eubacteria zymomonas mobilis (z. mobilis tgt) and escherichia coli (e. coli tgt) show a different behaviour in the presence of competitive inhibitors. the active sites of both enzymes are identical apart from a single conservative amino acid exchange, namely tyr106 of z. mobilis tgt is replaced by a phe in e. coli tgt. altho ...200314523925
altered glucose transport and shikimate pathway product yields in e. coli.different glucose transport systems are examined for their impact on phosphoenolpyruvate availability as reflected by the yields of 3-dehydroshikimic acid and byproducts 3-deoxy-d-arabino-heptulosonic acid, 3-dehydroquinic acid, and gallic acid synthesized by escherichia coli from glucose. 3-dehydroshikimic acid is an advanced shikimate pathway intermediate in the syntheses of a spectrum of commodity, pseudocommodity, and fine chemicals. all constructs carried plasmid arof(fbr) and tkta inserts ...200314524706
the glucanases of cellulomonas.cellulomonas is a unique bacterium possessing not only the capacity to degrade various carbohydrates, such as starch, xylan and cellulose, but crystalline cellulose as well. it has developed a complex battery of glucanases to deal with substrates possessing such extensive microheterogeneities. some of these enzymes are multifunctional, as well as cross inducible, possessing a multi-domain structure; these enzymes are thought to have arisen by the shuffling of these domains. intergeneric hybrids ...199714538714
zymomonas mobilis: a bacterium for ethanol production. 198614545375
the enzymatic hydrolysis and fermentation of pretreated wood substrates.aspenwood chips were pretreated by steam explosion. the various wood fractions obtained were assayed for their ability to act as substrates for growth and cellulase production of different trichoderma and clostridium thermocellum species. steam exploded aspenwood was as efficiently utilized as solka floc and correspondingly high cellulase activities were detected in the various culture filtrates. when t. harzianum e58 was grown on increasing concentrations of solka floc, highest cellulase and xy ...198414545694
genetic modification of zymomonas mobilis.the bacterium zymomonas mobilis is a potentially useful organism for the commercial production of ethanol as it is capable of more than double the rate of alcohol production by yeast. however, industrial application of this bacterium has been restricted in part due to the disadvantages of its limited substrate range (glucose, fructose and sucrose) and by-product formation. progress in strain improvement and genetic manipulation of this ethanologen is reviewed. methodologies for gaining reproduci ...199014549608
a novel and simple method for the purification of extracellular levansucrase from zymomonas mobilis.a new and simple method for the purification of extracellular levansucrase from zymomonas mobilis from highly viscous fermentation broth was developed. after incubation of the fermentation broth with a fructose-polymer cleaving enzyme preparation (fructozyme, novozymes, dk) for 48 h, levansucrase precipitated as aggregates and was redissolved in a 3 m urea solution. by ongoing size-exclusion chromatography on sephacryl s-300 the final levansucrase preparation was purified 100-fold and exhibited ...200314570269
characterization of native and histidine-tagged deoxyxylulose 5-phosphate reductoisomerase from the cyanobacterium synechocystis sp. pcc6803.the dxr gene encoding the 1-deoxy-d-xylulose 5-phosphate reductoisomerase (dxr) from the cyanobacterium synechocystis sp. pcc6803 was expressed in escherichia coli to produce both the native and n-terminal histidine-tagged forms of dxr. the enzymes were purified from the cell extracts using either anion exchange chromatography or metal affinity chromatography and gel filtration. the purified recombinant native and histidine-tagged enzymes each displayed a single band on sodium dodecyl sulfate-po ...200314580998
serine substitution for cysteine residues in levansucrase selectively abolishes levan forming activity.levansucrase is responsible for levan formation during sucrose fermentation of zymomonas mobilis, and this decreases the efficiency of ethanol production. as thiol modifying agents decrease levan formation, a role for cysteine residues in levansucrase activity has been examined using derivatives of z. mobilis levansucrase that carry serine substitutions of cysteine at positions 121, 151 or 244. these substitutions abolished the levan forming activity of levansucrase whilst only halving its activ ...200314584923
metabolic engineering of escherichia coli: construction of an efficient biocatalyst for d-mannitol formation in a whole-cell biotransformation.a whole-cell biotransformation system for the conversion of d-fructose to d-mannitol was developed in escherichia coli by constructing a recombinant oxidation/reduction cycle. first, the mdh gene, encoding mannitol dehydrogenase of leuconostoc pseudomesenteroides atcc 12291 (mdh), was expressed, effecting strong catalytic activity of an nadh-dependent reduction of d-fructose to d-mannitol in cell extracts of the recombinant e. coli strain. by contrast whole cells of the strain were unable to pro ...200414586579
disruption of the zymomonas mobilis extracellular sucrase gene (sacc) improves levan production.disruption of the extracellular zymomonas mobilis sucrase gene (sacc) to improve levan production.200415012804
crystallographic study of inhibitors of trna-guanine transglycosylase suggests a new structure-based pharmacophore for virtual screening.the enzyme trna-guanine transglycosylase (tgt) is involved in the pathogenicity of shigellae. as the crystal structure of this protein is known, it is a putative target for the structure-based design of inhibitors. here we report a crystallographic study of several new ligands exhibiting a 2,6-diamino-3h-quinazolin-4-one scaffold, which has been shown recently to be a promising template for tgt-inhibitors. crystal structure analysis of these complexes has revealed an unexpected movement of the s ...200415050823
performance of a newly developed integrant of zymomonas mobilis for ethanol production on corn stover hydrolysate.efficient conversion of lignocellulosic biomass requires biocatalysts able to tolerate inhibitors produced by many pretreatment processes. recombinant zymomonas mobilis 8b, a recently developed integrant of zymomonas mobilis 31821(pzb5), tolerated acetic acid up to 16 g l(-1) and achieved 82%-87% (w/w) ethanol yields from pure glucose/xylose solutions at ph 6 and temperatures of 30 degrees c and 37 degrees c. an ethanol yield of 85% (w/w) was achieved on glucose/xylose from hydrolysate produced ...200415055769
crystal structure of 1-deoxy-d-xylulose-5-phosphate reductoisomerase from zymomonas mobilis at 1.9-a resolution.1-deoxy-d-xylulose-5-phosphate reductoisomerase (dxr) is the second enzyme in the non-mevalonate pathway of isoprenoid biosynthesis. the structure of the apo-form of this enzyme from zymomonas mobilis has been solved and refined to 1.9-a resolution, and that of a binary complex with the co-substrate nadph to 2.7-a resolution. the subunit of dxr consists of three domains. residues 1-150 form the nadph binding domain, which is a variant of the typical dinucleotide-binding fold. the second domain c ...200415063313
biphasic aqueous/organic biotransformation of acetaldehyde and benzaldehyde by zymomonas mobilis pyruvate decarboxylase.zymomonas mobilis pyruvate decarboxylase (pdc) transformed acetaldehyde and benzaldehyde into (r)-phenylacetylcarbinol (pac), the precursor for the synthesis of ephedrine and pseudoephedrine. organic solvents were screened for a biphasic biotransformation with the enzyme in an aqueous phase and the toxic substrates delivered through the organic phase. in the absence of substrates a second phase of 1-pentanol, hexadecane or mtbe (methyl tertiary-butyl ether) stabilized the pdc activity in compari ...200415162454
antitumor activity of levan polysaccharides from selected microorganisms.levans were isolated from the cultures of gluconoacetobacter xylinus (g-levan; mw = 40,000), microbacterium laevaniformans (m; mw = 710,000), rahnella aquatilis (r; mw = 380,000), and zymomonas mobilis (z; mw = 570,000). the levans were composed mainly of fructose residues when analyzed by tlc and hplc, and their main backbones were beta-(2,6)-linkages with beta-(2,1)-branches by gc-ms and nmr. in the in vitro antitumor activity test of the levans against eight different tumor cell lines, relati ...200415178007
inhibition of thiamin diphosphate dependent enzymes by 3-deazathiamin diphosphate.3-deazathiamin diphosphate (deazatpp) and a second thiamin diphosphate (tpp) analogue having a benzene ring in place of the thiazolium ring have been synthesised. these compounds are both extremely potent inhibitors of pyruvate decarboxylase from zymomonas mobilis; binding is competitive with tpp and is essentially irreversible even though no covalent linkage is formed. deazatpp binds approximately seven-fold faster than tpp and at least 25,000-fold more tightly (k(i) less than 14 pm). deazatpp ...200415188040
use of zymomonas mobilis and saccharomyces cerevisiae mixed with kluyveromyces fragilis for improved ethanol production from jerusalem artichoke tubers.jerusalem artichoke mashed tubers were fermented using single yeasts and a bacterium as well as mixed culture of microorganisms. kluyveromyces fragilis, a yeast with an active inulinase, was used together with either a commercial distillery yeast, saccharomyces cerevisiae, or the bacterium zymomonas mobilis. after batch fermentation the best ethanol concentration of 0.48 g g(-1) for the mixed population and 0.46 g g(-1) for the single population can be obtained. the theoretical yield of the mixe ...200415269559
metabolic engineering of escherichia coli: construction of an efficient biocatalyst for d-mannitol formation in a whole-cell biotransformation.a whole-cell biotransformation system for the conversion of d-fructose to d-mannitol was developed in escherichia coli by construction of a recombinant oxidation/reduction cycle. first, the mdh gene encoding for the mannitol dehydrogenase of leuconostoc pseudomesenteroides atcc 12291 (mdh) was expressed, effecting a strong catalytic activity of a nadh-dependent reduction of d-fructose to d-mannitol in cell extracts of the recombinant e. coli strain but not enabling whole cells of the strain to p ...200315296170
catalytic activity of zymomonas mobilis extracellular "levan-levansucrase" complex in sucrose medium.the fructan biosynthesis by ethanol sedimented "levan-levansucrase" complex from zymomonas mobilis fermentation broth as well as purified levansucrase was investigated. the fructooligosaccharide (fos) producing activity of "levan-levamsucrase" sediment was investigated in 55% sucrose syrup at 45 degrees c. it was shown that fos in the syrup were presented by 1-kestose, 6-kestose, neokestose and nystose. the increase of gluconic acid concentration was observed in the reaction mixture during the i ...200315296187
crystallization and preliminary x-ray diffraction studies of a cobalt-substituted derivative of the iron-dependent alcohol dehydrogenase from zymomonas mobilis.the iron-dependent alcohol dehydrogenase from zymomonas mobilis has been crystallized in a form suitable for x-ray diffraction studies. the crystals grew in hanging drops by vapor diffusion, equilibrating with a solution comprising 25-27% methoxypolyethylene glycol 5000 and 1 mm co(2+) in a 0.2 m succinic acid/potassium hydroxide buffer at ph 5.5-5.7 at 281 k. crystals are tetragonal, p4(1)22 (or p4(3)22), with unit-cell dimensions a = b = 125.7, c = 248.1 a. four molecules comprise the asymmetr ...199615299751
inhibition of ethanol-producing yeast and bacteria by degradation products produced during pre-treatment of overview of the different inhibitors formed by pre-treatment of lignocellulosic materials and their inhibition of ethanol production in yeast and bacteria is given. different high temperature physical pre-treatment methods are available to render the carbohydrates in lignocellulose accessible for ethanol fermentation. the resulting hydrolyzsates contain substances inhibitory to fermentation-depending on both the raw material (biomass) and the pre-treatment applied. an overview of the inhibito ...200415300416
simultaneous saccharification and cofermentation of dilute-acid pretreated yellow poplar hardwood to ethanol using xylose-fermenting zymomonas mobilis.simultaneous saccharification and cofermentation (sscf) was carried out at approximately 15% total solids using conditioned dilute-acid pretreated yellow poplar feedstock, an adapted variant of national renewable energy laboratory (nrel) xylose-fermenting zymomonas mobilis and either commercial or nrel-produced cellulase enzyme preparations. in 7 d, at a cellulase loading of 12 filter paper units per gram cellulose (fpu/g), the integrated system produced more than 3% w/v ethanol and achieved 54% ...199915304685
ethanol production from corn starch in a fluidized-bed bioreactor.the production of ethanol from industrial dry-milled corn starch was studied in a laboratory-scale fluidized-bed bioreactor using immobilized biocatalysts. saccharification and fermentation were carried out either simultaneously or separately. simultaneous saccharification and fermentation (ssf) experiments were performed using small, uniform kappa-carrageenan beads (1.5-2.5 mm in diameter) of co-immobilized glucoamylase and zymomonas mobilis. dextrin feeds obtained by the hydrolysis of 15% drym ...199915304707
sorbitol can be produced not only chemically but also biotechnologically.sorbitol, a polyol found in many fruits, is attracting increasing industrial interest as a sweetener, humectant, texturizer, and softener. it is principally produced by chemical means. the bacterium zymomonas mobilis is able to produce sorbitol together with gluconic acid from fructose and glucose, respectively. this is possible in a one-step reaction via the enzyme glucose-fructose oxidoreductase, so far only known from z. mobilis. the possibilities for the production of sorbitol by z. mobilis ...200415304760
novel xylose dehydrogenase in the halophilic archaeon haloarcula marismortui.during growth of the halophilic archaeon haloarcula marismortui on d-xylose, a specific d-xylose dehydrogenase was induced. the enzyme was purified to homogeneity. it constitutes a homotetramer of about 175 kda and catalyzed the oxidation of xylose with both nadp+ and nad+ as cosubstrates with 10-fold higher affinity for nadp+. in addition to d-xylose, d-ribose was oxidized at similar kinetic constants, whereas d-glucose was used with about 70-fold lower catalytic efficiency (kcat/km). with the ...200415342590
parameter oscillations in a very high gravity medium continuous ethanol fermentation and their attenuation on a multistage packed column bioreactor system.the quasi-steady-states, marked by small fluctuations of residual glucose, ethanol, and biomass concentrations, and sustainable oscillations marked by big fluctuations of these monitored fermentation parameters were observed during the continuous ethanol fermentation of saccharomyces cerevisiae when very high gravity media were fed and correspondingly high ethanol concentrations reached. a high ethanol concentration was shown to be one of the main factors that incited these oscillations, althoug ...200415470717
the genome sequence of the ethanologenic bacterium zymomonas mobilis zm4.we report the complete genome sequence of zymomonas mobilis zm4 (atcc31821), an ethanologenic microorganism of interest for the production of fuel ethanol. the genome consists of 2,056,416 base pairs forming a circular chromosome with 1,998 open reading frames (orfs) and three ribosomal rna transcription units. the genome lacks recognizable genes for 6-phosphofructokinase, an essential enzyme in the embden-meyerhof-parnas pathway, and for two enzymes in the tricarboxylic acid cycle, the 2-oxoglu ...200415592456
ethanol fermentation on the move. 200515637618
zymomonas mobilis subspecies identification by amplified ribosomal dna restriction identify strains of zymomonas mobilis at the subspecies level by a fast and reliable technique.200515644116
high-level acetaldehyde production in lactococcus lactis by metabolic engineering.efficient conversion of glucose to acetaldehyde is achieved by nisin-controlled overexpression of zymomonas mobilis pyruvate decarboxylase (pdc) and lactococcus lactis nadh oxidase (nox) in l. lactis. in resting cells, almost 50% of the glucose consumed could be redirected towards acetaldehyde by combined overexpression of pdc and nox under anaerobic conditions.200515691976
experimental identification and quantification of glucose metabolism in seven bacterial species.the structurally conserved and ubiquitous pathways of central carbon metabolism provide building blocks and cofactors for the biosynthesis of cellular macromolecules. the relative uses of pathways and reactions, however, vary widely among species and depend upon conditions, and some are not used at all. here we identify the network topology of glucose metabolism and its in vivo operation by quantification of intracellular carbon fluxes from 13c tracer experiments. specifically, we investigated a ...200515716428
over-expression of xylulokinase in a xylose-metabolising recombinant strain of zymomonas mobilis.the broad host range vector pbbr1mcs-2 has been evaluated as an expression vector for zymomonas mobilis. the transformation efficiency of this vector was 2 x 10(3) cfu per mug of dna in a recombinant strain of z. mobilis zm4/acr containing the plasmid pzb5. stable replication for this expression vector was demonstrated for 50 generations. this vector was used to study xylose metabolism in acetate resistant z. mobilis zm4/acr (pzb5) by over-expression of xylulokinase (xk), as previous studies had ...200515727825
ethanol production from cellulosic materials by genetically engineered zymomonas confer the ability to ferment cello-oligosaccharides on the ethanol-producing bacterium, zymomonas mobilis, the beta-glucosidase gene from ruminococcus albus, tagged at its n-terminal with the 53-amino acid tat signal peptide from the periplasmic enzyme glucose-fructose oxidoreductase from z. mobilis, was introduced into the strain. the tag enabled 61% of the beta-glucosidase activity to be transported through the cytoplasmic membrane of the recombinant strain which then produced 0.49 g ethan ...200515742147
genome update: base skews in 200+ bacterial chromosomes. 200515758208
the effect of zymomonas mobilis culture on experimental schistosoma mansoni infection.c57bl/10 male mice infected with schistosoma mansoni were distributed into mixed, prophylactic and curative groups. a culture of zymomonas mobilis was orally administered to mice. a 61% protection from the infection was observed in the curative group (p <0.05). histopathological study of the livers and intestines showed similar results.200415765603
intermediates and transition states in thiamin diphosphate-dependent decarboxylases. a kinetic and nmr study on wild-type indolepyruvate decarboxylase and variants using indolepyruvate, benzoylformate, and pyruvate as substrates.the thiamin diphosphate (thdp)-dependent enzyme indolepyruvate decarboxylase (ipdc) is involved in the biosynthetic pathway of the phytohormone 3-indoleacetic acid and catalyzes the nonoxidative decarboxylation of 3-indolepyruvate to 3-indoleacetaldehyde and carbon dioxide. the steady-state distribution of covalent thdp intermediates of ipdc reacting with 3-indolepyruvate and the alternative substrates benzoylformate and pyruvate has been analyzed by (1)h nmr spectroscopy. for the first time, we ...200515835904
recombinant production of zymomonas mobilis pyruvate decarboxylase in the haloarchaeon haloferax volcanii.the unusual physiological properties of archaea (e.g., growth in extreme salt concentration, temperature and ph) make them ideal platforms for metabolic engineering. towards the ultimate goal of modifying an archaeon to produce bioethanol or other useful products, the pyruvate decarboxylase gene of zymomonas mobilis (zm pdc) was expressed in haloferax volcanii. this gene has been used successfully to channel pyruvate to ethanol in various gram-negative bacteria, including escherichia coli. altho ...200515876566
exchanging the substrate specificities of pyruvate decarboxylase from zymomonas mobilis and benzoylformate decarboxylase from pseudomonas putida.pyruvate decarboxylase from zymomonas mobilis (pdc) and benzoylformate decarboxylase from pseudomonas putida (bfd) are thiamine diphosphate-dependent enzymes that decarboxylate 2-keto acids. although they share a common homotetrameric structure they have relatively low sequence similarity and different substrate spectra. pdc prefers short aliphatic substrates whereas bfd favours aromatic 2-keto acids. these preferences are also reflected in their carboligation reactions. pdc catalyses the conver ...200515930043
the role of aspartic acid 143 in e. coli trna-guanine transglycosylase: insights from mutagenesis studies and computational modeling.trna guanine transglycosylase (tgt) is a trna-modifying enzyme which catalyzes the posttranscriptional exchange of guanine in position 34 of trna(y,h,n,d) with the modified base queuine in eukaryotes or its precursor, preq(1) base, in eubacteria. thus, tgt must recognize the guanine in trna and the free base queuine or preq(1) to catalyze this exchange. the crystal structure of zymomonas mobilis tgt with preq(1) bound suggests that a key aspartate is critically involved in substrate recognition. ...200515951383
kinetic control of thiamin diphosphate activation in enzymes studied by proton-nitrogen correlated nmr spectroscopy.proton-nitrogen correlated nmr studies were performed on thiamin diphosphate, which has been specifically labeled with (15)n at the 4'-amino group. after reconstitution of the labeled coenzyme with the apoenzymes of both wild-type pyruvate decarboxylase from zymomonas mobilis and the e50q variant, a high-field shift of the (15)n signal of approximately 4 ppm is observed at ph 5.9 when compared to that of the free coenzyme, indicating a higher electron density at the 4'-amino nitrogen in the enzy ...200515952776
[research progress of ethanologenic zymomonas mobilis].zymomonas mobilis is one of the natural ethanologenic microbes. with the unique entner-doudoroff pathway and some other special pathways of glycolytic and energetic metabolism, z. mobilis has remarkable characters of higher rate of ethanol production and higher tolerance to ethanol. glycolytic and energetic metabolism, tolerances (e.g., to ethanol, osmotic stress, etc.) and genetic improvements of z. mobilis are reviewed to elucidate the huge potential of z. mobilis in fuel ethanol production.200515989250
physiological regulation of the properties of alcohol dehydrogenase ii (adh ii) of zymomonas mobilis: nadh renders adh ii resistant to cyanide and aeration.the variable cyanide-sensitivity of the iron-containing alcohol dehydrogenase isoenzyme (adh ii) of the ethanol-producing bacterium zymomonas mobilis was studied. in aerobically grown permeabilized cells, cyanide caused gradual inhibition of adh ii, which was largely prevented by externally added nadh. cyanide-sensitivity of adh ii was highest in cells grown under conditions of vigorous aeration, in which intracellular nadh concentration was low. anaerobically grown bacteria, as well as those cu ...200516027951
transcriptional analysis of a gene cluster involved in glucose tolerance in zymomonas mobilis: evidence for an osmoregulated promoter.exponentially growing cells of zymomonas mobilis normally exhibit a lag period of up to 3 h when they are transferred from a liquid medium containing 2% glucose to a liquid medium containing 10% glucose. a mutant of z. mobilis (cu1) exhibited a lag period of more than 20 h when it was grown under the same conditions, whereas it failed to grow on a solid medium containing 10% glucose. the glucose-defective phenotype of mutant cu1 was due to a spontaneous insertion in a putative gene (orf4) identi ...200516030211
cog3926 and cog5526: a tale of two new lysozyme-like protein families.we have identified two new lysozyme-like protein families by using a combination of sequence similarity searches, domain architecture analysis, and structural predictions. first, the p5 protein from bacteriophage phi8, which belongs to cog3926 and pfam family duf847, is predicted to have a new lysozyme-like domain. this assignment is consistent with the lytic function of p5 proteins observed in several related double-stranded rna bacteriophages. domain architecture analysis reveals two lysozyme- ...200516155206
metabolic engineering of corynebacterium glutamicum for fuel ethanol production under oxygen-deprivation conditions.the central metabolic pathway of corynebacterium glutamicum was engineered to produce ethanol. a recombinant strain which expressed the zymomonas mobilis genes coding for pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adhb) was constructed. both genes placed under the control of the c. glutamicum ldha promoter were expressed at high levels in c. glutamicum, resulting, under oxygen-deprivation conditions, in a significant yield ofethanol from glucose in a process characterized by the abs ...200416179801
altered mrna expression of hepatic lipogenic enzyme and pparalpha in rats fed dietary levan from zymomonas mobilis.levan or high molecular beta-2,6-linked fructose polymer is produced extracellularly from sucrose-based substrates by bacterial levansucrase. in the present study, to investigate the effect of levan feeding on serum leptin, hepatic lipogenic enzyme and peroxisome proliferation-activated receptor (ppar) alpha expression in high-fat diet-induced obese rats, 4-week-old sprague-dawley male rats were fed high-fat diet (beef tallow, 40% of calories as fat), and, 6 weeks later, the rats were fed 0%, 1% ...200616214330
kinetic characterization of synechocystis sp. pcc6803 1-deoxy-d-xylulose 5-phosphate reductoisomerase mutants.the methylerythritol phosphate pathway to isoprenoids has been firmly established as an alternate to the mevalonate pathway in many bacteria, plants, algae, and the malaria parasite plasmodium falciparum. the second enzyme in this pathway, deoxy-d-xylulose 5-phosphate reductoisomerase (dxr; e.c., has been the focus of many investigations since it was found to be the target of the antibacterial and antimalarial compound, fosmidomycin. several x-ray crystal structures of the escherichia ...200616219495
isolation and characterization of mutants from levan-producing zymomonas mobilis.levan is a fructose polymer with potential importance as a fructose source and as a thickening agent in food technology. by n-methyl n'-nitro n-nitroso guanidine (ntg) mutagenesis and selection, two mutants, zml1 and zml2, of zymomonas mobilis b4286 overproducing levan (21.6 and 20.0 g/l at 24 h) were isolated. despite showing similar rates of sucrose hydrolysis (2.38 g/l/h) in fermentation, these mutants exhibited higher rates of levan production (1.72 and 1.68 g/l/h, respectively) than their p ...199916232453
investigation of the utility of pineapple juice and pineapple waste material as low-cost substrate for ethanol fermentation by zymomonas mobilis.the utility of the juice of rotten or discarded pineapples and the waste material of the production of pineapple juice as low-cost substrates for ethanol production by zymomonas mobilis was investigated. z. mobilis atcc 10988 produced 59.0 g.l(-1) ethanol in undiluted pineapple juice without nutritional supplementation and without the regulation of the ph while 42.5 g.l(-1) ethanol was obtained using a 125 g.l(-1) sucrose medium supplemented with 10 g.l(-1) yeast extract and mineral salts. ethan ...199916232532
optimization and interaction of media components in ethanol production using zymomonas mobilis by response surface methodology.ethanol production using zymomonas mobilis strains with medium optimized by response surface methodology (rsm) was studied. a significant improvement in the ethanol yield (0.50 g/g ethanol) was demonstrated, resulting in very low residual sugar and fewer by-products. the five-factor and five-response full factorial design have shown that glucose and yeast extract are the key media components that influence ethanol production, followed by the inoculum concentration and ammonium sulfate, with phos ...199916232623
production of ethanol from liquefied cassava starch using co-immobilized cells of zymomonas mobilis and saccharomyces cells of saccharomyces diastaticus and zymomonas mobilis produced a high ethanol concentration compared to immobilized cells of s. diastaticus during batch fermentation of liquefied cassava starch. the co-immobilized cells produced 46.7 g/l ethanol from 150 g/l liquefied cassava starch, while immobilized cells of yeast s. diastaticus produced 37.5 g/l ethanol. the concentration of ethanol produced by immobilized cells was higher than that by free cells of s. diastaticus and z. mob ...200116233146
high efficiency ethanol fermentation by entrapment of zymomonas mobilis into examine the potential of zymomonas mobilis entrapped into polyvinylalcohol (pva) lens-shaped immobilizates in batch and continuous ethanol production.200516238644
cofactor activation and substrate binding in pyruvate decarboxylase. insights into the reaction mechanism from molecular dynamics simulations.we have performed long-term molecular dynamics simulations of pyruvate decarboxylase from zymomonas mobilis. nine structures were modeled to investigate mechanistic questions related to binding of the cofactor, thiamin diphosphate (thdp), and the substrate in the active site. the simulations reveal that the proposed three thdp-tautomers all can bind in the active site and indicate that the equilibrium is shifted toward 4'-aminopyrimidine thdp in the absence of substrate. 4'-aminopyrimidinium thd ...200516274227
r-plasmid transfer in zymomonas mobilis.conjugal transfer of three incp1 plasmids and one incfii plasmid into strains of the ethanol-producing bacterium zymomonas mobilis was obtained. these plasmids were transferred at high frequencies from escherichia coli and pseudomonas aeruginosa into z. mobilis and also between different z. mobilis strains, using the membrane filter mating technique. most of the plasmids were stably maintained in z. mobilis, although there was some evidence of delayed marker expression. a low level of chromosoma ...198016345598
comparison of ethanol production by different zymomonas strains.a comparison of the rates of growth and ethanol production by 11 different strains of zymomonas revealed a wide range of characteristics, with some strains being more tolerant of high sugar or ethanol concentrations and high incubation temperatures than others. some strains were unable to utilize sucrose; others produced large amounts of levan, and one strain grew well but produced no levan. one strain, cp4, was considerably better in all respects than most of the other strains and was chosen as ...198116345753
controlling morphological instability of zymomonas mobilis strains in continuous culture.growth of zymomonas mobilis atcc 29191 and cp4 in a continuous stirred tank fermentor resulted in the selection of stable flocculating variants. factors responsible for enhancing the system pressures selective for the morphological variants were identified. by incorporating some modifications into the design of the fermentor, it was possible to achieve steady-state operation of the chemostat with both wild-type and flocculating strains. biochemical and microscopic studies were performed to eluci ...198316346320
controlling morphological instability of zymomonas mobilis strains in continuous culture.[this corrects the article on p. 1900 in vol. 45.].198316346393
expression of a lactose transposon (tn951) in zymomonas mobilis.the potential utility of zymomonas mobilis as an organism for the commercial production of ethanol would be greatly enhanced by the addition of foreign genes which expand its range of fermentable substrates. we tested various plasmids and mobilizing factors for their ability to act as vectors and introduce foreign genes into z. mobilis cp4. plasmid pgc91.14, a derivative of rp1, was found to be transferred from escherichia coli to z. mobilis at a higher frequency than previously reported for any ...198316346422
d-glucose transport system of zymomonas mobilis.the properties of the d-glucose transport system of zymomonas mobilis were determined by measuring the uptake of nonmetabolizable analogs (2-deoxy-d-glucose and d-xylose) by wild-type cells and the uptake of d-glucose itself by a mutant lacking glucokinase. d-glucose was transported by a constitutive, stereospecific, carrier-mediated facilitated diffusion system, whereby its intracellular concentration quickly reached a plateau close to but not above the external concentration. d-xylose was tran ...198516346694
isolation of noninhibitory strains of zymomonas mobilis.wild-type zymomonas mobilis strains inhibit the growth of escherichia coli. we report the first isolation of noninhibitory strains, called zymomonas inhibition negative (zin), after treatment with n-methyl-n'-nitro-n-nitrosoguanidine. a standardized soft-agar overlay procedure for detecting e. coli growth inhibition was also developed.198516346764
thermal tolerance of zymomonas mobilis: temperature-induced changes in membrane composition.the membrane composition of zymomonas mobilis changed dramatically in response to growth temperature. with increasing temperature, the proportion of vaccenic acid declined with an increase in myristic acid, the proportion of phosphatidylcholine and cardiolipin increased with decreases in phosphatidylethanolamine and phosphatidylglycerol, and the phospholipid/protein ratio of the membrane declined. these changes in membrane composition were correlated with changes in thermal tolerance and with ch ...198616347087
transfer of plasmids to an antibiotic-sensitive mutant of zymomonas mobilis.wild-type strains of zymomonas mobilis exhibit multiple antibiotic resistance and thus restrict the use of many broad-host-range plasmids in them as cloning vehicles. antibiotic-sensitive mutants of z. mobilis were isolated and used as hosts for the conjugal transfer of broad-host-range plasmids from escherichia coli. such antibiotic-sensitive strains can facilitate the application of broad-host-range plasmids to the study of z. mobilis.198616347136
expression vector for zymomonas mobilis.this study describes the construction of several useful cloning vectors which can be conjugated from escherichia coli into zymomonas mobilis at high frequency, approaching 10 per donor or recipient. these vectors contain a broad-host-range replicon and mob site from rsf1010, a chloramphenicol acyltransferase gene under the control of an enteric consensus promoter, and a second mob site (originally derived from rp4). the addition of this second mob site appears to be responsible for a 2-order-of- ...198716347272
zymomonas mobilis mutants with an increased rate of alcohol production.two new derivatives of zymomonas mobilis cp4 were isolated from enrichment cultures after 18 months of serial transfers. these new strains were selected for the ability to grow and produce ethanol rapidly on transfer into fresh broth containing ethanol and allyl alcohol. ethanol production by these strains was examined in batch fermentations under three sets of conditions. both new derivatives were found to be superior to the parent strain cp4 with respect to the speed and completeness of glucos ...198716347373
fermentation of d-xylose and l-arabinose to ethanol by erwinia chrysanthemi.erwinia spp. are gram-negative facultative anaerobes within the family enterobacteriacae which possess several desirable traits for the conversion of pentose sugars to ethanol, such as the ability to ferment a broad range of carbohydrates and the ease with which they can be genetically modified. twenty-eight strains of erwinia carotovora and e. chrysanthemi were screened for the ability to ferment d-xylose to ethanol. e. chrysanthemi b374 was chosen for further study on the basis of its superior ...198716347426
fermentation of d-xylose to ethanol by genetically modified klebsiella planticola.d-xylose is a plentiful pentose sugar derived from agricultural or forest residues. enteric bacteria such as klebsiella spp. ferment d-xylose to form mixed acids and butanediol in addition to ethanol. thus the ethanol yield is normally low. zymomonas spp. and most yeasts are unable to ferment xylose, but they do ferment hexose sugars to ethanol in high yield because they contain pyruvate decarboxylase (ec, a key enzyme that is absent from enteric bacteria. this report describes the ferm ...198716347427
production of acetaldehyde by zymomonas mobilis.mutants of zymomonas mobilis were selected for decreased alcohol dehydrogenase activity by using consecutively higher concentrations of allyl alcohol. a mutant selected by using 100 mm allyl alcohol produced acetaldehyde at a level of 4.08 g/liter when the organism was grown in aerated batch cultures on a medium containing 4.0% (wt/wt) glucose. on the basis of the amount of glucose utilized, this level of acetaldehyde production represents nearly 40% of the maximum theoretical yield. acetaldehyd ...198716347497
expression of different levels of ethanologenic enzymes from zymomonas mobilis in recombinant strains of escherichia coli.the expression of zymomonas mobilis genes encoding pyruvate decarboxylase and alcohol dehydrogenase ii in escherichia coli converted this organism from the production of organic acids to the production of ethanol. ethanol was produced during both anaerobic and aerobic growth. the extent to which these ethanologenic enzymes were expressed correlated with the extent of ethanol production. the replacement of organic acids with ethanol as a metabolic product during aerobic and anaerobic growth resul ...198816347553
microbiology of ripening honey.two main groups of bacteria, classified as gluconobacter and lactobacillus, are present in ripening honey. a third bacterial group, classified as zymomonas, and several types of yeast are occasionally isolated. both in natural honey and in synthetic syrup the bacterial population decreases in the course of the ripening process. lactobacillus and gluconobacter disappear after minimum moisture (about 18%) is reached, but the former does so sooner than the latter. the presence of these bacteria in ...197516350044
polyphasic study of zymomonas mobilis strains revealing the existence of a novel subspecies z. mobilis subsp. francensis subsp. nov., isolated from french cider.zymomonas mobilis strains recently isolated from french 'framboisé' ciders were compared with collection strains of the two defined subspecies, z. mobilis subsp. mobilis and z. mobilis subsp. pomaceae, using a polyphasic approach. six strains isolated from six different regions of france were compared with three strains of z. mobilis subsp. mobilis, including the type strain lmg 404t, and four strains of z. mobilis subsp. pomaceae, including the type strain lmg 448t, using phenotypic and genotyp ...200616403876
enhanced benzaldehyde tolerance in zymomonas mobilis biofilms and the potential of biofilm applications in fine-chemical production.biotransformation plays an increasingly important role in the industrial production of fine chemicals due to its high product specificity and low energy requirement. one challenge in biotransformation is the toxicity of substrates and/or products to biocatalytic microorganisms and enzymes. biofilms are known for their enhanced tolerance of hostile environments compared to planktonic free-living cells. zymomonas mobilis was used in this study as a model organism to examine the potential of surfac ...200616461720
tfam detects co-evolution of trna identity rules with lateral transfer of histidyl-trna synthetase.we present tfam, an automated, statistical method to classify the identity of trnas. tfam, currently optimized for bacteria, classifies initiator trnas and predicts the charging identity of both typical and atypical trnas such as suppressors with high confidence. we show statistical evidence for extensive variation in trna identity determinants among bacterial genomes due to variation in overall tdna base content. with tfam we have detected the first case of eukaryotic-like trna identity rules i ...200616473847
the ice nucleation gene from pseudomonas syringae as a sensitive gene reporter for promoter analysis in zymomonas mobilis.the expression of the ice nucleation gene inaz from pseudomonas syringae in zymomonas mobilis strains under the control of three different promoters was investigated to establish the utility of the gene as a reporter and examine the possible use of the organism as a source of ice nuclei for biotechnological applications. a promoterless version of the inaz gene was placed under the control of three different promoters: p(infpdc) (pyruvate decarboxylase), a homologous strong promoter from z. mobil ...199516534909
isolation and properties of mutants of zymomonas mobilis deficient in sugar enrichment method using d-cycloserine was designed for the isolation of spontaneous mutants of zymomonas mobilis deficient in glucose or fructose utilization. the mutants could easily be isolated since they represented 80 to 90% of the population after two and three enrichment cycles. glucokinase and fructokinase activities in the mutants were affected.199616535260
kinetics of sugar transport and phosphorylation influence glucose and fructose cometabolism by zymomonas mobilis.the competitive inhibition of fructokinase by glucose has been proposed as the mechanism by which zymomonas mobilis preferentially consumes glucose from mixtures of glucose and fructose and accumulates fructose when growing on sucrose. in this study, incorporation of radioactive fructose into biomass was used as a measure of fructose catabolism. it was determined that the rate of fructose incorporation by z. mobilis cp4 was somewhat lower in the presence of an equimolar concentration of glucose ...199716535690
cosmeceutical properties of levan produced by zymomonas mobilis.levan, a polysaccharide that can be produced by both plants and microorganisms, is a sugar polymer composed of fructose, with beta-2,6 linkages. here, we have attempted to assess the possible use of levan produced by zymomonas mobilis as a cosmeceutical ingredient. in service of this goal, we assessed a host of levan's properties, including its moisturizing effects, cell cytotoxicity, cell proliferation effects, and anti-inflammation effects. levan exhibited a moisturizing effect that was almost ...200516538295
kinetic modeling to optimize pentose fermentation in zymomonas mobilis.zymomonas mobilis engineered to express four heterologous enzymes required for xylose utilization ferments xylose along with glucose. a network of pentose phosphate (pp) pathway enzymatic reactions interacting with the native glycolytic entner doudoroff (ed) pathway has been hypothesized. we have investigated this putative reaction network by developing a kinetic model incorporating all of the enzymatic reactions of the pp and ed pathways, including those catalyzed by the heterologous enzymes. s ...200616570322
measurement and analysis of intracellular atp levels in metabolically engineered zymomonas mobilis fermenting glucose and xylose mixtures.intracellular adenosine-5'-triphosphate (atp) levels were measured in a metabolically engineered zymomonas mobilis over the course of batch fermentations of glucose and xylose mixtures. fermentations were conducted over a range of ph (5-6) in the presence of varying initial amounts of acetic acid (0-8 g/l) using a 10% (w/v) total sugar concentration (glucose only, xylose only, or 5% glucose/5% xylose mixture). over the design space investigated, ethanol process yields varied between 56.6% and 92 ...200616599547
the 1.9 a resolution structure of mycobacterium tuberculosis 1-deoxy-d-xylulose 5-phosphate reductoisomerase, a potential drug target.1-deoxy-d-xylulose 5-phosphate reductoisomerase catalyzes the nadph-dependent rearrangement and reduction of 1-deoxy-d-xylulose 5-phosphate to form 2-c-methyl-d-erythritol 4-phosphate, as the second step of the deoxyxylulose 5-phosphate/methylerythritol 4-phosphate pathway found in many bacteria and plants. the end product, isopentenyl diphosphate, is the precursor of various isoprenoids vital to all living organisms. the pathway is not found in humans; the mevalonate pathway is instead used for ...200616790937
enhancement of acid tolerance in zymomonas mobilis by a proton-buffering peptide.a portion of the cbpa gene from escherichia coli k-12 encoding a 24 amino acid proton-buffering peptide (pbp) was cloned via the shuttle vector pjb99 into e. coli jm105 and subsequently into zymomonas mobilis cp4. expression of pbp was confirmed in both jm105 and cp4 by hplc. z. mobilis cp4 carrying pjb99-2 (pbp) exhibited increased acid tolerance (p < 0.05) in acidified tsb (hcl [ph 3.0] or acetic acid [ph 3.5]), glycine-hcl buffer (ph 3.0), and sodium acetate-acetic acid buffer (ph 3.5) in com ...200616891663
crystal structure of nadp(h)-dependent 1,5-anhydro-d-fructose reductase from sinorhizobium morelense at 2.2 a resolution: construction of a nadh-accepting mutant and its application in rare sugar synthesis.recombinant 1,5-anhydro-d-fructose reductase (afr) from sinorhizobium morelense s-30.7.5 was crystallized in complex with the cofactor nadp(h) and its structure determined to 2.2 a resolution using selenomethionine sad (refined r(work) and r(free) factors of 18.9 and 25.0%, respectively). as predicted from the sequence and shown by the structure, afr can be assigned to the gfo/idh/moca protein family. afr consists of two domains. the n-terminal domain displays a rossmann fold and contains the co ...200616906761
zymomonas mobilis as catalyst for the biotechnological production of sorbitol and gluconic acid.the conversion of glucose and fructose into gluconic acid (ga) and sorbitol (sor) was conducted in a batch reactor with free (ctab-treated or not) or immobilized cells of zymomonas mobilis. high yields (more than 90%) of gluconic acid and sorbitol were attained at initial substrate concentration of 600 g/l (glucose plus fructose at 1:1 ratio), using cells with glucose-fructose-oxidoreductase activity of 75 u/l. the concentration of the products varied hyperbolically with time according to the eq ...200616915688
microdiesel: escherichia coli engineered for fuel production.biodiesel is an alternative energy source and a substitute for petroleum-based diesel fuel. it is produced from renewable biomass by transesterification of triacylglycerols from plant oils, yielding monoalkyl esters of long-chain fatty acids with short-chain alcohols such as fatty acid methyl esters and fatty acid ethyl esters (faees). despite numerous environmental benefits, a broader use of biodiesel is hampered by the extensive acreage required for sufficient production of oilseed crops. ther ...200616946248
respiratory behaviour of a zymomonas mobilis adhb::kan(r) mutant supports the hypothesis of two alcohol dehydrogenase isoenzymes catalysing opposite reactions.perturbation of the aerobic steady-state in a chemostat culture of the ethanol-producing bacterium zymomonas mobilis with a small pulse of ethanol causes a burst of ethanol oxidation, although the reactant ratio of the alcohol dehydrogenase (adh) reaction ([nadh][acetaldehyde][h(+)])/([ethanol][nad(+)]) remains above the k(eq) value. simultaneous catalysis of ethanol synthesis and oxidation by the two adh isoenzymes, residing in different redox microenvironments, has been proposed previously. in ...200616950260
ethanol production from dilute-acid softwood hydrolysate by co-culture.dilute-acid softwood hydrolysate, with glucose and xylose as the dominant sugars, was fermented to ethanol by co-cultures. the strains used include saccharomyces cerevisiae 2.535 (1#), pachysolen tannophilis atcc 2.1662 (2#), and recombinant escherichia coli (3#) constructed in our laboratory carrying both pdc and adhb genes derived from zymomonas mobilis. before fermentation, the co-cultures were adapted for five batches. observation under light microscope showed aggregation of adapted strains, ...200616960285
physiology of zymomonas mobilis: some unanswered questions.the ethanol-producing bacterium zymomonas mobilis can serve as a model organism for the study of rapid catabolism and inefficient energy conversion in bacteria. some basic aspects of its physiology still remain poorly understood. here, the energy-spilling pathways during uncoupled growth, the structure and function of electron transport chain, and the possible reasons for the inefficient oxidative phosphorylation are analysed. also, the interaction between ethanol synthesis and respiration is co ...200617010696
[progress on engineered strains for ethanol production].with the 21 century's coming, the era of cheap oil is coming to the end. there has been an increasing worldwide interest in fuel ethanol. in the last two decades, lots of work has been done to develop strains for ethanol producing. research progress on metabolic engineering of strains for fuel ethanol production is summarized, including genetically engineered saccharomyces cerevisiae to utilize starch, pentose and cellulose, zymomonas mobilis to ferment arabinose and xylose, escherichia coli and ...200617037078
bacterial diversity in the active stage of a bioremediation system for mineral oil hydrocarbon-contaminated soils.soils contaminated with mineral oil hydrocarbons are often cleaned in off-site bioremediation systems. in order to find out which bacteria are active during the degradation phase in such systems, the diversity of the active microflora in a degrading soil remediation system was investigated by small-subunit (ssu) rrna analysis. two sequential rna extracts from one soil sample were generated by a procedure incorporating bead beating. both extracts were analysed separately by generating individual ...200617074900
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