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probing the location and function of the conserved histidine residue of phosphoglucose isomerase by using an active site directed inhibitor n-bromoacetylethanolamine phosphate.phosphoglucose isomerase (ec 5.3.1.9) catalyzes the interconversion of d-glucopyranose-6-phosphate and d-fructofuranose-6-phosphate by promoting an intrahydrogen transfer between c1 and c2. a conserved histidine exists throughout all phosphoglucose isomerases and was hypothesized to be the base catalyzing the isomerization reaction. in the present study, this conserved histidine, his311, of the enzyme from bacillus stearothermophilus was subjected to mutational analysis, and the mutational effec ...199910595547
effects of acetate on the growth and fermentation performance of escherichia coli ko11.escherichia coli ko11, in which the genes pdc (pyruvate decarboxylase) and adh (alcohol dehydrogenase) encoding the ethanol pathway from zymomonas mobilis were inserted into the chromosome, has been shown to metabolize all major sugars that are constituents of hemicellulosic hydrolysates to ethanol, in anaerobic conditions. however, the growth and fermentation performance of this recombinant bacteria may be affected by acetic acid, a potential inhibitor present in hemicellulose hydrolysates in a ...199910652785
pulsed high-field gradient in vivo nmr spectroscopy to measure diffusional water permeability in corynebacterium glutamicum.pulsed high-field gradient in vivo nmr spectroscopy was used to measure diffusional water permeability in cell suspensions of the gram-positive bacterium corynebacterium glutamicum. two different regions of h2o mobility were detected. one was characterized by the apparent coefficient of self-diffusion, d(1 app) = (4.6-12.7)x10(-8) cm(2) s(-1), depending on the observation time t. the other region was characterized by d(2) = 1.4x10(-5) cm(2) s(-1). the value of d(2) was similar to the diffusion c ...200010683237
refolding and purification of zymomonas mobilis levansucrase produced as inclusion bodies in fed-batch culture of recombinant escherichia coli.zymomonas mobilis levansucrase was overproduced by the fed-batch culture of recombinant escherichia coli harboring a novel expression system that is constitutively expressed by the promoter from the rahnella aquatilis levansucrase gene. most of the levansucrase was produced as inclusion bodies in the bacterial cytoplasm, accounting for approximately 20% of the total cellular protein. refolding after complete denaturation by high concentrations of urea or guanidine hydrochloride was not successfu ...200010733894
the effect of osmo-induced stress on product formation by zymomonas mobilis on sucrose.the intensification of biosynthesis of fructooligosaccharides in the presence of high salt concentrations was observed during sucrose (10%) fermentation by zymomonas mobilis 113s. a 0.6 m nacl concentration led to an increase of oligosaccharide productivity by 3.5-fold. sorbitol formation was increased in the presence of 0.16 m nacl and was inhibited at highest salt concentrations. in a medium with high (65%, w/w) sucrose content the salts gave inhibitory effects on fructooligosaccharide product ...200010791734
symbiotic induction of pyruvate dehydrogenase genes from sinorhizobium meliloti.genes coding for components of the pyruvate dehydrogenase (pdh) multienzyme complex (pdhc) from sinorhizobium meliloti, the alfalfa symbiont, have been isolated on the basis of their high expression in symbiotic bacteria. the elp component, pdh, is encoded by two genes, pdhaalpha (1,047 bp) and pdhabeta (1,383 bp), a situation encountered in the alpha-proteobacteria rickettsia prowazekii and zymomonas mobilis as well as in some gram-positive bacteria and in mitochondria. pdhaalpha and pdhabeta p ...200010796014
cyanide inhibits respiration yet stimulates aerobic growth of zymomonas mobilis.potassium cyanide at submillimolar concentrations (20-500 microm) inhibited the high respiration rates of aerobic cultures of zymomonas mobilis but, remarkably, stimulated culture growth. in batch culture, after an extended lag phase, exponential growth persisted longer, resulting in higher biomass densities. in aerobic chemostat cultures, elevated biomass concentration was observed in the presence of cyanide. this growth stimulation effect is attributed to decreased production of the inhibitory ...200010846205
simultaneous saccharification and cofermentation of peracetic acid-pretreated biomass.previous work in our laboratories has demonstrated the effectiveness of peracetic acid for improving enzymatic digestibility of lignocellulosic materials. the use of dilute alkali solutions as a pre-pretreatment prior to peracetic acid lignin oxidation increased carbohydrate hydrolysis yields in a synergistic as opposed to additive manner. deacetylation of xylan is easily achieved using dilute alkali solutions under mild conditions. in this article, we evaluate the effectiveness of peracetic aci ...200010849783
development of new ethanologenic escherichia coli strains for fermentation of lignocellulosic biomass.two new ethanologenic strains (fbr4 and fbr5) of escherichia coli were constructed and used to ferment corn fiber hydrolysate. the strains carry the plasmid ploi297, which contains the genes from zymomonas mobilis necessary for efficiently converting pyruvate into ethanol. both strains selectively maintained the plasmid when grown anaerobically. each culture was serially transferred 10 times in anaerobic culture with sugar-limited medium containing xylose, but no selective antibiotic. an average ...200010849788
comparative energetics of glucose and xylose metabolism in recombinant zymomonas mobilis.recombinant zymomonas mobilis cp4:pzb5 was grown with ph control in batch and continuous modes with either glucose or xylose as the sole carbon and energy source. in batch cultures in which the ratio of the final cell mass concentration to the amount of sugar in the medium was constant (i.e., under conditions that promote "coupled growth"), maximum specific rates of glucose and xylose consumption were 8.5 and 2.1 g/(g of cell.h), respectively; maximum specific rates of ethanol production for glu ...200010849796
continuous fermentation studies with xylos-utilizing recombinant zymomonas mobilis.this study examined the continuous cofermentation performance characteristics of a dilute-acid "prehydrolysate-adapted" recombinant zymomonas 39676:pzb4l and builds on the ph-stat batch fermentations with this recombinant that we reported on last year. substitution of yeast extract by 1% (w/v) corn steep liquor (csl) (50% solids) and mg (2 mm) did not alter the cofermentation performance. using declared assumptions, the cost of using csl and mg was estimated to be 12.5 cents/gal of ethanol with ...200010849797
characterization of a high-productivity recombinant strain of zymomonas mobilis for ethanol production from glucose/xylose mixtures.the fermentation characteristics of a recombinant strain of zymomonas mobilis zm4(pzb5) capable of converting both glucose and xylose to ethanol have been further investigated. previous studies have shown that the strain zm4(pzb5) was capable of converting a mixture of 65 g/l of glucose and 65 g/l of xylose to 62 g/l of ethanol in 48 h with an overall yield of 0.46 g/g. higher sugar concentrations (e.g., 75/75 g/l) resulted in incomplete xylose utilization (80 h). in the present study, further k ...200010849801
nuclear magnetic resonance studies of acetic acid inhibition of rec zymomonas mobilis zm4(pzb5).the fermentation characteristics and effects of lignocellulosic toxic compounds on recombinant zymomonas mobilis zm4(pzb5), which is capable of converting both glucose and xylose to ethanol, and its parental strain, zm4, were characterized using 13c and 31p nuclear magnetic resonance (nmr) in vivo. from the 31p nmr data, the levels of nucleoside triphosphates (ntp) of zm(pzb5) using xylose were lower than those of glucose. this can be related to the intrinsically slower assimilation and/or metab ...200010849802
ethanol production from glucose and xylose by immobilized zymomonas mobilis cp4(pzb5).fermentation of glucose-xylose mixtures to ethanol was investigated in batch and continuous experiments using immobilized recombinant zymomonas mobilis cp4(pzb5). this microorganism was immobilized by entrapment in kappa-carrageenan beads having a diameter of 1.5-2.5 mm. batch experiments showed that the immobilized cells cofermented glucose and xylose to ethanol and that the presence of glucose improved the xylose utilization rate. batch fermentation of rice straw hydrolysate containing 76 g/l ...200010849817
the effect of overliming on the toxicity of dilute acid pretreated lignocellulosics: the role of inorganics, uronic acids and ether-soluble organics.although the treatment of dilute acid pretreated lignocellulosics with calcium hydroxide or carbonate (overliming) is known to improve the fermentability of carbohydrate-rich hydrolyzate streams, a firm understanding of the chemistry behind the process is lacking. quantitative evaluation of inorganics, uronic acids, and non-polar organics indicates that only a portion of the improvement can be ascribed to these materials. upon overliming the concentrations of inorganics either increase (ca, mg), ...200010899549
molecular weight and antitumour activity of zymomonas mobilis levans.levans produced by several zymomonas mobilis strains were classified by their viscosity average molecular weight and tested against sarcoma 180. measurements of the samples' polydispersity were carried out to characterise the molecular weight distribution. the antitumour activities of levan samples were plotted against the viscosity average molecular weight and a maximum value of this activity was found at mv = 210,000 (mw = 456,900 and pd = 16.2). the results indicate that levan antitumour acti ...200010921850
effects of deletions at the carboxyl terminus of zymomonas mobilis pyruvate decarboxylase on the kinetic properties and substrate specificity.the three-dimensional structure of zymomonas mobilis pyruvate decarboxylase shows that the carboxyl-terminal region of the protein occludes the active site. this observation is consistent with earlier suggestions that the active site is inaccessible to solvent during catalysis. however, the carboxyl-terminal region must move aside to allow entry of the substrate, and again to permit the products to leave. here we have examined the role of the carboxyl terminus by making 15 variants of the enzyme ...200010924138
characterization and replication properties of the zymomonas mobilis atcc 10988 plasmids pzmo1 and pzmo2.the complete nucleotide sequences of two small cryptic zymomonas mobilis atcc 10988 plasmids (pzmo1 and pzmo2) were determined. the plasmids showed 67% homology to each other at their nucleotide level. plasmid pzmo1 was 1651 bp long with 38% g + c content and contained an open reading frame (orfzmo1) of 1044 nucleotides. orfzmo1 is predicted to encode a polypeptide of 348 amino acids and shows a high degree of homology with gram-negative replication proteins of rolling circle replicating plasmid ...200010964623
identification of heptadecanoic and c19 cyclopropane fatty acids in the lipid fraction of zymomonas mobilis.in the present paper the presence of heptadecanoic acid and c19 cyclopropane fatty acid in zymomonas mobilis is reported. the former has not been mentioned as a component of the lipid fraction of this microorganism, and the presence of the latter is not well documented.200010977901
proposal of sphingomonadaceae fam. nov., consisting of sphingomonas yabuuchi et al. 1990, erythrobacter shiba and shimidu 1982, erythromicrobium yurkov et al. 1994, porphyrobacter fuerst et al. 1993, zymomonas kluyver and van niel 1936, and sandaracinobacter yurkov et al. 1997, with the type genus sphingomonas yabuuchi et al. 1990.based on the results of phylogenetic analysis of the 16s rdna sequences and the presence of n-2'-hydroxymyristoyl dihydrosphingosine 1-glucuronic acid (sgl-1) and 2-hydroxymyristic acid (non-hydroxymyristic acid in zymomonas) in cellular lipids, a new family, sphingomonadaceae, for group 4 of the alpha-subclass of the class proteobacteria is herein proposed and a description of the family is given. the family consists of six genera, sphingomonas, erythrobacter, erythromicrobium, porphyrobacter, ...200010981829
rapid detection of zymomonas mobilis redox activity using 5-cyano-2,3-tolyl-tetrazolium chloride (ctc). 200010997252
cloning and sequencing of the levansucrase gene from acetobacter xylinum nci 1005.the levansucrase gene (lsxa) was cloned from the genomic dna of acetobacter xylinum nci 1005, and the nucleotide sequence of the lsxa gene (1,293 bp) was determined. the deduced amino acid sequence of the lsxa gene showed 57.4% and 46.2% identity with the levansucrases from zymomonas mobilis and erwinia amylovora, respectively, while only 35.2% identity with that from acetobacter diazotrophicus. the gene product of lsxa (lsxa) that was overproduced in e. coli coded for a polypeptide of molecular ...200010997873
isolation of the dxr gene of zymomonas mobilis and characterization of the 1-deoxy-d-xylulose 5-phosphate reductoisomerase.the gene encoding the second enzyme of the 2c-methyl-d-erythritol 4-phosphate (mep) pathway for isopentenyl diphosphate biosynthesis, 1-deoxy-d-xylulose 5-phosphate (dxp) reductoisomerase, was cloned and sequenced from zymomonas mobilis. the deduced amino acid sequence showed the highest identity (48.2%) to the dxp reductoisomerase of escherichia coli. biochemical characterization of the purified dxp reductoisomerase showed a strict dependence of the enzyme on nadph and divalent cations (mn(2+), ...200011004410
mutagenesis at asp27 of pyruvate decarboxylase from zymomonas mobilis. effect on its ability to form acetoin and acetolactate.pyruvate decarboxylase (pdc) is one of several enzymes that require thiamin diphosphate (thdp) and a bivalent cation as essential cofactors. the three-dimensional structure of pdc from zymomonas mobilis (zmpdc) shows that asp27 (d27) is close to thdp in the active site, and mutagenesis of this residue has suggested that it participates in catalysis. the normal product of the pdc reaction is acetaldehyde but it is known that the enzyme can also form acetoin as a by-product from the hydroxyethyl-t ...200011029594
sorbitol and gluconic acid production using permeabilized zymomonas mobilis cells confined by hollow-fiber membranes.immobilization of zymomonas mobilis by different methods was investigated. experiments were performed in order to choose the most appropriate support for the immobilization of the cells. the most advantageous option was to use permeabilized cells in the bore of microporous hollow fibers. whereas the reaction rate was about 33 g of gluconate/(g of protein x h) using hollow fibers, which is comparable to that observed by using free cells, the calcium alginate immobilized cells presented a reaction ...200011069007
effect of feeding strategy on zymomonas mobilis cp4 fed-batch fermentations and mathematical modeling of the system.in this work, the effect of the feeding strategy in zymomonas mobilis cp4 fed-batch fermentations on the final biomass and ethanol concentrations was studied. highest glucose yields to biomass (0.018 g/g) and to ethanol (0.188 g/g) were obtained in fed-batch fermentations carried out using different feeding rates with a glucose concentration in the feed equal to 100 g/l. lower values (0.0102 g biomass/g glucose and 0.085 g ethanol/ g glucose) were obtained when glucose accumulated to levels high ...200011092622
crystal structure of a truncated mutant of glucose-fructose oxidoreductase shows that an n-terminal arm controls tetramer formation.n-terminal or c-terminal arms that extend from folded protein domains can play a critical role in quaternary structure and other intermolecular associations and/or in controlling biological activity. we have tested the role of an extended n-terminal arm in the structure and function of a periplasmic enzyme glucose-fructose oxidoreductase (gfor) from zymomonas mobilis. we have determined the crystal structure of the nad(+) complex of a truncated form of the enzyme, gfordelta, in which the first 2 ...200011099381
influence of the flocculating agent in sedimentation and performance of a non flocculent strain of zymomonas mobilis in the ethanol production process.the influence of the flocculating agent was studied in the performance (measured by microbial growth and ethanol production) of a non flocculent strain of zymomonas mobilis, as well as the potentiality of the sedimentation process in the separation of the biomass from the fermentation broth. among the flocculating agents studied, it was verified that both tannin and the polyelectrolyte yielded good results with regard to cellular performance. however, with regard to sedimentation tannin is more ...200011105240
simultaneous ethanol and bacterial ice nuclei production from sugar beet molasses by a zymomonas mobilis cp4 mutant expressing the inaz gene of pseudomonas syringae in continuous culture.the aim of this work was to construct a zymomonas mobilis mutant capable of simultaneous ethanol and ice nuclei production from agricultural by-product such as sugar beet molasses, in steady-state continuous culture.200011123473
crystal versus solution structures of thiamine diphosphate-dependent enzymes.the quaternary structures of the thiamine diphosphate-dependent enzymes transketolase (ec 2.2.1.1; from saccharomyces cerevisiae), pyruvate oxidase (ec 1.2.3.3; from lactobacillus plantarum), and pyruvate decarboxylase (ec 4.1.1.1; from zymomonas mobilis and brewers' yeast, the latter in the native and pyruvamide-activated forms) were examined by synchrotron x-ray solution scattering. the experimental scattering data were compared with the curves calculated from the crystallographic models of th ...200010617618
kinetic and nuclear magnetic resonance studies of xylose metabolism by recombinant zymomonas mobilis zm4(pzb5).the specific rates of growth, substrate utilization, and ethanol production as well as yields of biomass and ethanol production on xylose for the recombinant zymomonas mobilis zm4(pzb5) were shown to be much less than those on glucose or glucose-xylose mixtures. typical fermentations with zm4(pzb5) growing on glucose-xylose mixtures followed two-phase growth kinetics with the initial uptakes of glucose and xylose being followed by slower growth and metabolic uncoupling on xylose after glucose de ...200010618222
molecular characterization of squalene synthase from the green microalga botryococcus braunii, race b.the green microalga botryococcus braunii produces large amounts of liquid hydrocarbons and is classified into three races, depending on the type of the hydrocarbon produced. the b race produces two types of triterpenoid hydrocarbons, squalene and botryococcene, both of which are putative condensation products of farnesyl diphosphate. in an attempt to better understand the regulation involved in the production of squalene and botryococcene, we have isolated and characterized a squalene synthase ( ...200010620354
construction and characterization of an effector strain of streptococcus mutans for replacement therapy of dental caries.an effector strain has been constructed for use in the replacement therapy of dental caries. recombinant dna methods were used to make the streptococcus mutans supercolonizing strain, jh1140, lactate dehydrogenase deficient by deleting virtually all of the ldh open reading frame (orf). to compensate for the resulting metabolic imbalance, a supplemental alcohol dehydrogenase activity was introduced by substituting the adhb orf from zymomonas mobilis in place of the deleted ldh orf. the resulting ...200010639415
altered regulation of pyruvate kinase or co-overexpression of phosphofructokinase increases glycolytic fluxes in resting escherichia coli.glycolytic fluxes in resting escherichia coli were enhanced by overexpression of heterologous pyruvate kinases (pyk) from bacillus stearothermophilus and zymomonas mobilis, but not homologous pyk. compared to the control, an increase of 10% in specific glucose consumption and of 15% in specific ethanol production rates was found in anaerobic resting cells, expressing the heterologous pyks, that were harvested from exponentially growing aerobic cultures. a further increase in glycolytic flux was ...200010649237
gene integration and expression and extracellular secretion of erwinia chrysanthemi endoglucanase cely (cely) and celz (celz) in ethanologenic klebsiella oxytoca p2.the development of methods to reduce costs associated with the solubilization of cellulose is essential for the utilization of lignocellulose as a renewable feedstock for fuels and chemicals. one promising approach is the genetic engineering of ethanol-producing microorganisms that also produce cellulase enzymes during fermentation. by starting with an ethanologenic derivative (strain p2) of klebsiella oxytoca m5a1 with the native ability to metabolize cellobiose, the need for supplemental beta- ...200111133422
specificity of signal peptide recognition in tat-dependent bacterial protein translocation.the bacterial twin arginine translocation (tat) pathway translocates across the cytoplasmic membrane folded proteins which, in most cases, contain a tightly bound cofactor. specific amino-terminal signal peptides that exhibit a conserved amino acid consensus motif, s/t-r-r-x-f-l-k, direct these proteins to the tat translocon. the glucose-fructose oxidoreductase (gfor) of zymomonas mobilis is a periplasmic enzyme with tightly bound nadp as a cofactor. it is synthesized as a cytoplasmic precursor ...200111133954
methanogen and bacterial diversity and distribution in deep gas hydrate sediments from the cascadia margin as revealed by 16s rrna molecular analysis.the microbial community of a deep (to 234 m below the sea floor) sediment gas hydrate deposit (cascadia margin ocean drilling program site 889/890, leg 146) was analysed for the first time by molecular genetic techniques. both bacterial and methanogen diversity were determined by phylogenetic analysis of ribosomal dna sequences. high molecular mass dna, indicative of active bacteria, was present in all of the samples. ribosomal rna genes were amplified from extracted dna extracted from sediment ...200111137602
on-line monitoring and controlling system for fermentation processes.a personal computer-based on-line monitoring and controlling system was developed for the fermentation of microorganism. the on-line hplc system for the analysis of glucose and ethanol in the fermentation broth was connected to the fermenter via an auto-sampling equipment, which could perform the pipetting, filtration and dilution of the sample and final injection onto the hplc through automation based on a programmed procedure. the a/d and d/a interfaces were equipped in order to process the si ...200111150792
a new target for shigellosis: rational design and crystallographic studies of inhibitors of trna-guanine transglycosylase.eubacterial trna-guanine transglycosylase (tgt) is involved in the hyper-modification of cognate trnas leading to the exchange of g34 at the wobble position in the anticodon loop by preq1 (2-amino-5-(aminomethyl)pyrrolo[2,3-d]pyrimidin-4(3h)-one) as part of the biosynthesis of queuine (q). mutation of the tgt gene in shigella flexneri results in a significant loss of pathogenicity of the bacterium, revealing tgt as a new target for the design of potent drugs against shigellosis. the x-ray struct ...200111178905
molecular organization of the ribosomal rna transcription unit and the phylogenetic study of zymomonas mobilis zm4.previously we reported that zymomonas mobilis zm4 contains three ribosomal transcription units (rrna to c operons) which are clustered around the 50 min region, paci fragments 13 and 6, on the physical map of z. mobilis zm4 [kang, h. l. and kang, h. s. (1998) gene 206, 223-228]. the physical map reveals that the rrna gene set is located on the 76 kb paci fragment 13. the complete nucleotide sequence of the rrna gene set has been determined. the total number of nucleotides of the rrna gene set is ...200111266123
engineering a homo-ethanol pathway in escherichia coli: increased glycolytic flux and levels of expression of glycolytic genes during xylose fermentation.replacement of the native fermentation pathway in escherichia coli b with a homo-ethanol pathway from zymomonas mobilis (pdc and adhb genes) resulted in a 30 to 50% increase in growth rate and glycolytic flux during the anaerobic fermentation of xylose. gene array analysis was used as a tool to investigate differences in expression levels for the 30 genes involved in xylose catabolism in the parent (strain b) and the engineered strain (ko11). of the 4,290 total open reading frames, only 8% were ...200111325924
consequences of a modified putative substrate-activation site on catalysis by yeast pyruvate decarboxylase.earlier, it had been proposed in the laboratories at halle that a cysteine residue is responsible for the hysteretic substrate activation behavior of yeast pyruvate decarboxylase. more recently, this idea has received support in a series of studies from rutgers with the identification of residue c221 as the site where substrate is bound to transmit the information to h92, to e91, to w412, and finally to the active center thiamin diphosphate. according to steady-state kinetic assays, the c221a/c2 ...200111327837
structural diversity of the triterpenic hydrocarbons from the bacterium zymomonas mobilis: the signature of defective squalene cyclization by the squalene/hopene cyclase.twelve polycyclic triterpenic hydrocarbons (alpha- and gamma-polypodatetraenes, dammara-20(21),24-diene, 17-isodammara-12,24-diene, eupha-7,24-diene, hop-17(21)-ene, neohop-13(18)-ene, 17-isodammara-20(21),24-diene, neohop-12-ene, fern-8-ene, diploptene and hop-21-ene) were detected in the hydrocarbon fraction from the bacterium zymomonas mobilis. some of them have never been reported from bacteria. these triterpenes were present in z. mobilis in significant amounts, comparable to those of diplo ...200111377875
a simple and efficient method for the purification of membrane-bound levansucrase from zymomonas mobilis.a new and efficient method for the purification of levansucrase from cell-free extracts of a flocculant mutant of zymomonas mobilis atcc 10988 was developed. levansucrase activity was almost completely recovered and purified by a factor of 15 after precipitation with 0.1 m mncl2 as a first capturing step. the enzyme was homogeneously purified by ultrafiltration and anion-exchange chromatography and exhibited a levan-forming activity of 39.2 u mg-1. the native enzyme formed large aggregates with ...200111381333
production of glucose-fructose oxidoreductase and ethanol by zymomonas mobilis atcc 29191 in medium containing corn steep liquor as a source of vitamins.different concentrations of corn steep liquor (csl) were tested in the cultivation of zymomonas mobilis. cell growth, ethanol production, and the formation of glucose-fructose oxidoreductase (gfor) and glucono-delta-lactonase (gl), the enzymes responsible for the bio-production of gluconic acid and sorbitol, were examined. the cell yields using 25 g csl l(-1) and 40 g csl l(-1) (y(x,s) approximately 0.031 g g(-1)) were close to that obtained with 5 g yeast extract (ye) l(-1). with 5 g csl l(-1) ...200111398924
iszm1068: an is5-like insertion element from zymomonas mobilis.a new insertion sequence, designated iszm1068, was isolated from zymomonas mobilis strain cp4. this element consists of 1,068 bp and contains one major orf which shows similarities both at the nucleotide and at the amino acid sequence level with the corresponding orfs encoding the transposases of many is5 family elements, in particular the is1031 group. moreover, the z. mobilis orf shares the conserved n2, n3 and c1 signature motifs of the is4 and is5 families. six out of seven z. mobilis wild-t ...200111409542
continuous production of (r)-phenylacetylcarbinol in an enzyme-membrane reactor using a potent mutant of pyruvate decarboxylase from zymomonas mobilis.the optimization of a continuous enzymatic reaction yielding (r)-phenylacetylcarbinol (pac), an intermediate of the l-ephedrine synthesis, is presented. we compare the suitability of three pyruvate decarboxylases (pdc), pdc from saccharomyces cerevisiae, pdc from zymomonas mobilis, and a potent mutant of the latter, pdcw392m, with respect to their application in the biotransformation using acetaldehyde and benzaldehyde as substrates. among these, the mutant enzyme was the most active and most st ...200111410856
site-directed mutagenesis of the ionizable groups in the active site of zymomonas mobilis pyruvate decarboxylase: effect on activity and ph dependence.pyruvate decarboxylase (pdc, ec 4.1.1.1) is a thiamin diphosphate-dependent enzyme about which there is a large body of structural and functional information. the active site contains several absolutely conserved ionizable groups and all of these appear to be important, as judged by the fact that mutation diminishes or abolishes catalytic activity. previously we have shown [schenk, g., leeper, f.j., england, r., nixon, p.f. & duggleby, r.g. (1997) eur. j. biochem. 248, 63-71] that the activity i ...200111422387
use of a green fluorescent protein gene as a reporter in zymomonas mobilis and halomonas elongata.we investigated the applicability of the green fluorescent protein of aequorea victoria as a reporter for gene expression in the strictly fermentative gram-negative ethanologenic bacterium zymomonas mobilis and in the moderately halophilic bacterium halomonas elongata. we have succeeded to express a mutated gene of green fluorescent protein under the control of different promoters in z. mobilis and h. elongata grown under various glucose or salt concentrations, respectively. our results demonstr ...200111470365
bifurcation analysis of continuous biochemical reactor models.the validity of a biochemical reactor model often is evaluated by comparing transient responses to experimental data. dynamic simulation can be a rather inefficient and ineffective tool for analyzing bioreactor models that exhibit complex nonlinear behavior. bifurcation analysis is a powerful tool for obtaining a more efficient and complete characterization of the model behavior. to illustrate the power of bifurcation analysis, the steady-state and transient behavior of three continuous bioreact ...200111485425
fuel ethanol production from lignocellulose: a challenge for metabolic engineering and process integration.with industrial development growing rapidly, there is a need for environmentally sustainable energy sources. bioethanol (ethanol from biomass) is an attractive, sustainable energy source to fuel transportation. based on the premise that fuel bioethanol can contribute to a cleaner environment and with the implementation of environmental protection laws in many countries, demand for this fuel is increasing. efficient ethanol production processes and cheap substrates are needed. current ethanol pro ...200111499926
crystal structure of histidinol phosphate aminotransferase (hisc) from escherichia coli, and its covalent complex with pyridoxal-5'-phosphate and l-histidinol phosphate.the biosynthesis of histidine is a central metabolic process in organisms ranging from bacteria to yeast and plants. the seventh step in the synthesis of histidine within eubacteria is carried out by a pyridoxal-5'-phosphate (plp)-dependent l-histidinol phosphate aminotransferase (hisc, ec 2.6.1.9). here, we report the crystal structure of l-histidinol phosphate aminotransferase from escherichia coli, as a complex with pyridoxamine-5'-phosphate (pmp) at 1.5 a resolution, as the internal aldimine ...200111518529
production of the gram-positive sarcina ventriculi pyruvate decarboxylase in escherichia coli.sarcina ventriculi grows in a remarkable range of mesophilic environments from ph 2 to ph 10. during growth in acidic environments, where acetate is toxic, expression of pyruvate decarboxylase (pdc) serves to direct the flow of pyruvate into ethanol during fermentation. pdc is rare in bacteria and absent in animals, although it is widely distributed in the plant kingdom. the pdc gene from s. ventriculi is the first to be cloned and characterized from a gram-positive bacterium. in escherichia col ...200111535783
the chryseobacterium meningosepticum pafa enzyme: prototype of a new enzyme family of prokaryotic phosphate-irrepressible alkaline phosphatases?chryseobacterium meningosepticum is an aerobic gram-negative rod widely distributed in natural environments. unlike many bacteria, it produces a phosphate-irrepressible periplasmic alkaline phosphatase (ap). this work describes cloning of the gene encoding that enzyme from c. meningosepticum ccug 4310 (nctc 10585), and preliminary characterization of its product. the gene, named pafa, encodes a protein (pafa) of 546 amino acids with a calculated molecular mass of the mature peptide of 58682 da. ...200111577161
crystal structures of the precursor form of glucose-fructose oxidoreductase from zymomonas mobilis and its complexes with bound ligands.the nadp(h)-dependent enzyme glucose-fructose oxidoreductase (gfor) is a classic example of a redox protein that is translocated across a membrane in fully folded form. gfor is synthesized in the cytoplasm with a 52-residue signal peptide, giving a precursor form, pregfor, that is fully active and has its cofactor tightly bound. a twin-arginine motif in the signal peptide directs it to a sec-independent pathway by which it is translocated, in fully folded form, into the periplasm where it functi ...200111705375
trna-guanine transglycosylase from escherichia coli: molecular mechanism and role of aspartate 89.the enzyme trna-guanine transglycosylase (tgt, ec 2.4.2.29) catalyzes a posttranscriptional transglycosylation reaction involved in the incorporation of the modified base queuine [q, 7-(4,5-cis-dihydroxy-2-cyclopenten-1-ylaminomethyl)-7-deazaguanine] into trna. previously, the crystal structure of the tgt from zymomonas mobilis was solved in complex with preq(1) (the substrate for the eubacterial tgt) [romier et al. (1996) embo j. 15, 2850-2857]. an aspartate residue at position 102 (position 89 ...200111714265
pyruvate decarboxylase: a key enzyme for the oxidative metabolism of lactic acid by acetobacter pasteurianus.acetobacter pasteurianus, an obligately oxidative bacterium, is the first organism shown to utilize pyruvate decarboxylase (pdc) as a central enzyme for oxidative metabolism. in plants, yeast, and other bacteria, pdc functions solely as part of the fermentative ethanol pathway. during the growth of a. pasteurianus on lactic acid, the central intermediate pyruvate is cleaved to acetaldehyde and co(2) by pdc. acetaldehyde is subsequently oxidized to its final product, acetic acid. the presence of ...200111734888
zymomonas mobilis cp4 fed-batch fermentations of glucose-fructose mixtures to ethanol and sorbitol.zymomonas mobilis cp4 fed-batch fermentations of glucose-fructose mixtures were carried out at different operational conditions (aeration, feed rate and substrate concentration) to test their effects on the system productivity. in these fermentations, the main products were ethanol and sorbitol. kinetic parameters were calculated using the experimental data. however, parameters in the sorbitol synthesis rate were estimated from data recorded in different experiments in order to avoid the effect ...200111759679
production of ethanol from liquefied cassava starch using co-immobilized cells of zymomonas mobilis and saccharomyces diastaticus.co-immobilized 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
fermentation performance assessment of a genomically integrated xylose-utilizing recombinant of zymomonas mobilis 39676.in ph-controlled batch fermentations with pure sugar synthetic hardwood hemicellulose (1% [w/v] glucose and 4% xylose) and corn stover hydrolysate (8% glucose and 3.5% xylose) lacking acetic acid, the xylose-utilizing, tetracycline (tc)-sensitive, genomically integrated variant of zymomonas mobilis atcc 39676 (designated strain c25) exhibited growth and fermentation performance that was inferior to national renewable energy laboratory's first-generation, tc-resistant, plasmid-bearing zymomonas r ...200111963841
comparative ethanol productivities of different zymomonas recombinants fermenting oat hull hydrolysate.iogen corporation of ottawa, canada, has recently built a 50 t/d biomass-to-ethanol demonstration plant adjacent to its enzyme production facility. iogen has partnered with the university of toronto to test the c6/c5 cofermentation performance characteristics of national renewable energy laboratory's metabolically engineered zymomonas mobilis using its biomass hydrolysates. in this study, the biomass feedstock was an agricultural waste, namely oat hulls, which was hydrolyzed in a proprietary two ...200111963842
enzymes of the entner-doudoroff and pyruvate decarboxylation pathways in zymomonas mobilis wild-type cp4 and mutant strains grown in continuous culture.the osmotolerant zymomonas mobilis strain suc40, (containing plasmid pds3154-inaz), which is capable of producing simultaneously ethanol and ice nuclei protein, was cultivated in a chemically defined complete sucrose medium, as well as in a sugar beet molasses medium in continuous culture. the strain exhibited the normal monod's relationship between biomass and dilution rate, and between growth substrate concentration and dilution rate. specific activities of a number of enzymes that appear to c ...200111827214
aerobic and facultatively anaerobic cellulolytic bacteria from the gut of the termite zootermopsis angusticollis.to demonstrate the occurrence of cellulolytic bacteria in the termite zootermopsis angusticollis.200211849325
flux through citrate synthase limits the growth of ethanologenic escherichia coli ko11 during xylose fermentation.previous studies have shown that high levels of complex nutrients (luria broth or 5% corn steep liquor) were necessary for rapid ethanol production by the ethanologenic strain escherichia coli ko11. although this strain is prototrophic, cell density and ethanol production remained low in mineral salts media (10% xylose) unless complex nutrients were added. the basis for this nutrient requirement was identified as a regulatory problem created by metabolic engineering of an ethanol pathway. cells ...200211872452
de novo design, synthesis, and in vitro evaluation of inhibitors for prokaryotic trna-guanine transglycosylase: a dramatic sulfur effect on binding affinity. 200211921407
purification of food-grade oligosaccharides using immobilised cells of zymomonas mobilis.immobilised cells of the bacterium zymomonas mobilis were used to remove glucose, fructose, and sucrose from food-grade oligosaccharide mixtures. unpurified fructo-, malto-, isomalto-, gentio-, and inulinoligosaccharides, containing total carbohydrate concentrations of 300 g l(-1), were added to immobilised cells, in 100 ml batch reactors. no ph control or nutrient additions were required. contaminating glucose, fructose, and sucrose within the mixtures was completely fermented within 12 h. the ...200211935179
semicontinuous production of ethanol from agricultural wastes by immobilised coculture in a two stage bioreactor.the seed testing laboratories of maharashtra discard 10 tonnes of grains and oil seeds treated with pesticides per annum. these agricultural wastes could be converted to reducing sugar and ethanol in a two stage semicontinuous fluidised bed bioreactor containing immobilised, bacillus sp. and zymomonas mobilis in the 1st stage and saccharomyces diastaticus and s. cerevisae in the 2nd stage. the optimum temperature and ph for fermentation in both the stages were 30 degrees c and 7.2 respectively. ...200212674381
steady-state measurements of lactic acid production in a wild-type and a putative d-lactic acid dehydrogenase-negative mutant of zymomonas mobilis: influence of glycolytic flux.this work represents a continuation of our investigation into environmental conditions that promote lactic acid synthesis by zymomonas mobilis. the characteristic near theoretical yield of ethanol from glucose by z. mobilis can be compromised by the synthesis of d- and l-lactic acid. the production of lactic acid is exacerbated by the following conditions: ph 6.0, yeast extract, and reduced growth rate. at a specific growth rate of 0.048/h, the average yield of dl-lactate from glucose in a yeast ...200212018249
characterization of heterologous and native enzyme activity profiles in metabolically engineered zymomonas mobilis strains during batch fermentation of glucose and xylose mixtures.zymomonas mobilis has been metabolically engineered to broaden its substrate utilization range to include d-xylose and l-arabinose. both genomically integrated and plasmid-bearing z. mobilis strains that are capable of fermenting the pentose d-xylose have been created by incorporating four genes: two genes encoding xylose utilization metabolic enzymes (xyla/xylb) and two genes encoding pentose phosphate pathway enzymes (talb/tkta). we have characterized the activities of the four newly introduce ...200212018261
performance testing of zymomonas mobilis metabolically engineered for cofermentation of glucose, xylose, and arabinose.iogen corporation of ottawa, canada, has recently built a 40t/d biomass-to-ethanol demonstration plant adjacent to its enzyme production facility. it has partnered with the university of toronto to test the c6/c5 cofermenta-tion performance characteristics of the national renewable energy labora-tory's metabolically engineered zymomonas mobilis using various biomass hydrolysates. iogen's feedstocks are primarily agricultural wastes such as corn stover and wheat straw. integrated recombinant z. m ...200212018270
cofermentation of glucose, xylose, and arabinose by genomic dna-integrated xylose/arabinose fermenting strain of zymomonas mobilis ax101.cofermentation of glucose, xylose, and arabinose is critical for complete bioconversion of lignocellulosic biomass, such as agricultural residues and herbaceous energy crops, to ethanol. we have previously developed a plasmid-bearing strain of zymomonas mobilis (206c[pzb301]) capable of cofermenting glucose, xylose, and arabinose to ethanol. to enhance its genetic stability, several genomic dna-integrated strains of z. mobilis have been developed through the insertion of all seven genes necessay ...200212018310
performance of immobilized zymomonas mobilis 31821 (pzb5) on actual hydrolysates produced by arkenol technology.by applying the arkenol process using highly concentrated sulfuric acid, various biomass feedstocks, including cedar tree, rice straw, newspaper, and bagasse, were successfully processed and converted into glucose and xylose for fermentation usage in a flash fermentation reactor in which the performance of national renewable energy laboratory's patented rec-zymomonas mobilis 31821 (pzb5) after immobilization was investigated. the immobilization medium is a photocrosslinked resin made from polyet ...200212018312
evaluation of a recombinant klebsiella oxytoca strain for ethanol production from cellulose by simultaneous saccharification and fermentation: comparison with native cellobiose-utilising yeast strains and performance in co-culture with thermotolerant yeast and zymomonas mobilis.in the simultaneous saccharification and fermentation to ethanol of 100 g l(-1) microcrystalline cellulose, the cellobiose-fermenting recombinant klebsiella oxytoca p2 outperformed a range of cellobiose-fermenting yeasts used in earlier work, despite producing less ethanol than reported earlier for this organism under similar conditions. the time taken by k. oxytoca p2 to produce up to about 33 g l(-1) ethanol was much less than for any other organism investigated, including ethanol-tolerant str ...200212039532
cloning and characterization of the zymobacter palmae pyruvate decarboxylase gene (pdc) and comparison to bacterial homologues.pyruvate decarboxylase (pdc) is the key enzyme in all homo-ethanol fermentations. although widely distributed among plants, yeasts, and fungi, pdc is absent in animals and rare in bacteria (established for only three organisms). genes encoding the three known bacterial pdc genes have been previously described and expressed as active recombinant proteins. the pdc gene from zymomonas mobilis has been used to engineer ethanol-producing biocatalysts for use in industry. in this paper, we describe a ...200212039744
modeling and advanced control of recombinant zymomonas mobilis fed-batch fermentation.this work presents the development of an unstructured kinetic model incorporating the differing degrees of product, substrate, and ph inhibition on the kinetic rates of ethanol fermentation by recombinant zymomonas mobilis cp4:pzb5 for growth on two substrates. product inhibition was observed to start affecting the specific growth rate at an ethanol concentration of 20 g/l and the specific productivity at about 35-40 g/l. specific growth rate was also shown to be more sensitive to inhibition by ...200212052075
respective effects of sodium and chloride ions on filament formation and growth and ethanol production in zymomonas mobilis fermentations.to elucidate the separate effects of the cation and anion in zymomonas mobilis cultures inhibited by nacl.200212081545
ethanol cycle in an ethanologenic bacterium.a novel redox cycle is suggested, performing interconversion between acetaldehyde and ethanol in aerobically growing ethanologenic bacterium zymomonas mobilis. it is formed by the two alcohol dehydrogenase (adh) isoenzymes simultaneously catalyzing opposite reactions. adh i is catalyzing acetaldehyde reduction. the local reactant ratio at its active site probably is shifted towards ethanol synthesis due to direct channeling of nadh from glycolysis. adh ii is oxidizing ethanol. the net result of ...200212095609
the biotechnological production of sorbitol.sorbitol, a polyol found in many fruits, is of increasing industrial interest as a sweetener, humectant, texturizer and softener. at present, it is produced chemically. the bacterium zymomonas mobilis is able to produce sorbitol and gluconic acid from fructose and glucose, respectively. this is possible in a one-step reaction via a glucose-fructose oxidoreductase so far only known from z. mobilis. the possibilities for the industrial production of sorbitol by z. mobilis are discussed, and compar ...200212172602
interplay of sos induction, recombinant gene expression, and multimerization of plasmid vectors in escherichia coli.using pbr322- and puc-derived plasmid vectors, a homologous (escherichia coli native esterase) and three heterologous proteins (human interleukin-2, human interleukin-6, and zymomonas levansucrase) were synthesized in e. coli ic2015(reca::lacz) and gy4786 (sfia::lacz) strains. via time-course measurement of beta-galactosidase activity in each recombinant culture, the sos induction was estimated in detail and the results were systematically compared. in recombinant e. coli, the sos response did n ...200212209789
identification of functionally important amino acid residues in zymomonas mobilis levansucrase.the catalytic residues of levansucrase (sucrose:2,6-beta-d-fructan 6-beta-d-fructosyltransferase, ec 2.4.1.10) from zymomonas mobilis were analyzed by random mutation and site-directed mutagenesis. we found that substitution of glu278 with asp and his reduced the k(cat) for sucrose hydrolysis 30- and 210-fold, respectively, strongly suggesting glu278 plays a key role in catalyzing this reaction. given the likelihood that another acidic amino residue was also involved, we constructed variants in ...200212359071
improvement of cellulolytic properties of clostridium cellulolyticum by metabolic engineering.cellulolytic clostridia have evolved to catabolize lignocellulosic materials at a seasonal biorhythm, so their biotechnological exploitation requires genetic improvements. as high carbon flux leads to pyruvate accumulation, which is responsible for the cessation of growth of clostridium cellulolyticum, this accumulation is decreased by heterologous expression of pyruvate decarboxylase and alcohol dehydrogenase from zymomonas mobilis. in comparison with that of the wild strain, growth of the reco ...200211772608
solvent kinetic isotope effects monitor changes in hydrogen bonding at the active center of yeast pyruvate decarboxylase concomitant with substrate activation: the substituent at position 221 can control the state of activation.substrate activation of yeast pyruvate decarboxylase has been studied extensively in the authors' laboratories providing strong evidence that interaction of substrate with residue c221 provides the trigger, and the information is then transmitted along the c221 to h92 to e91 to w412 to g413 pathway to the 4'-amino nitrogen of the thiamin diphosphate cofactor. earlier, it was found that the c221s substitution reduced the hill coefficient from 2.0 to 0.8-0.9, the c221a substitution to 1.0, even th ...200211781083
levan fructotransferase from arthrobacter oxydans j17-21 catalyzes the formation of the di-d-fructose dianhydride iv from levan.a new levan fructotransferase (lftase) isolated from arthrobacter oxydans j17-21 was characterized for the production of difructose dianhydride iv (dfa iv). lftase was purified to apparent homogeneity by q-sepharose ion exchange chromatography, mono-q hr 5/5 column chromatography, and gel permeation chromatography. the enzyme had an apparent molecular mass of 54000 da. the optimum ph for the enzyme-catalyzed reaction was ph 6.5, and the optimum temperature was observed at 45 degrees c. the lftas ...200312696949
composite hopanoid biosynthesis in zymomonas mobilis: n-acetyl-d-glucosamine as precursor for the cyclopentane ring linked to bacteriohopanetetrol.a selective labelling of the two major composite hopanoids of z. mobilis with deuterated n-acetyl-d-glucosamine showed that this carbohydrate is a common precursor of the glucosamine or the cyclopentitol moieties respectively linked to bacteriohopanetetrol by a glycosidic or an ether bond.200312703821
cellulosic fuel ethanol: alternative fermentation process designs with wild-type and recombinant zymomonas mobilis.iogen (canada) is a major manufacturer of industrial cellulase and hemicellulase enzymes for the textile, pulp and paper, and poultry feed industries. iogen has recently constructed a 40 t/d biomass-to-ethanol demonstration plant adjacent to its enzyme production facility. the integration of enzyme and ethanol plants results in significant reduction in production costs and offers an alternative use for the sugars generated during biomass conversion. iogen has partnered with the university of tor ...200312721468
engineering lactic acid bacteria with pyruvate decarboxylase and alcohol dehydrogenase genes for ethanol production from zymomonas mobilis.lactic acid bacteria are candidates for engineered production of ethanol from biomass because they are food-grade microorganisms that can, in many cases, metabolize a variety of sugars and grow under harsh conditions. in an effort to divert fermentation from production of lactic acid to ethanol, plasmids were constructed to express pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adh), encoded by the pdc and adhb genes of zymomonas mobilis, in lactic acid bacteria. several strains were tr ...200312750944
crystal structure of thiamindiphosphate-dependent indolepyruvate decarboxylase from enterobacter cloacae, an enzyme involved in the biosynthesis of the plant hormone indole-3-acetic acid.the thiamin diphosphate-dependent enzyme indolepyruvate decarboxylase catalyses the formation of indoleacetaldehyde from indolepyruvate, one step in the indolepyruvate pathway of biosynthesis of the plant hormone indole-3-acetic acid. the crystal structure of this enzyme from enterobacter cloacae has been determined at 2.65 a resolution and refined to a crystallographic r-factor of 20.5% (rfree 23.6%). the subunit of indolepyruvate decarboxylase contains three domains of open alpha/beta topology ...200312752451
studies on structure-function relationships of indolepyruvate decarboxylase from enterobacter cloacae, a key enzyme of the indole acetic acid pathway.enterobacter cloacae, isolated from the rhizosphere of cucumbers, produces large amounts of indole-3-acetic acid. indolepyruvate decarboxylase, the key enzyme in the biosynthetic pathway of indole-3-acetic acid, catalyses the formation of indole-3-acetaldehyde and carbon dioxide from indole-3-pyruvic acid. the enzyme requires the cofactors thiamine diphosphate and magnesium ions for catalytic activity. recombinant indolepyruvate decarboxylase was purified from the host escherichia coli strain jm ...200312752452
phosphoenolpyruvate availability and the biosynthesis of shikimic acid.the impact of increased availability of phosphoenolpyruvate during shikimic acid biosynthesis has been examined in escherichia coli k-12 constructs carrying plasmid-localized arof(fbr) and tkta inserts encoding, respectively, feedback-insensitive 3-deoxy-d-arabino-heptulosonic acid 7-phosphate synthase and transketolase. strategies for increasing the availability of phosphoenolpyruvate were based on amplified expression of e. coli ppsa-encoded phosphoenolpyruvate synthase or heterologous express ...200312790643
detection of aromatic alpha-hydroxyketones with tetrazolium salts. 200312824586
nmr analysis of covalent intermediates in thiamin diphosphate enzymes.enzymic catalysis proceeds via intermediates formed in the course of substrate conversion. here, we directly detect key intermediates in thiamin diphosphate (thdp)-dependent enzymes during catalysis using (1)h nmr spectroscopy. the quantitative analysis of the relative intermediate concentrations allows the determination of the microscopic rate constants of individual catalytic steps. as demonstrated for pyruvate decarboxylase (pdc), this method, in combination with site-directed mutagenesis, en ...200312834340
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 products.in 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
virtual screening for submicromolar leads of trna-guanine transglycosylase based on a new unexpected binding mode detected by crystal structure analysis.eubacterial trna-guanine transglycosylase (tgt) is involved in the hypermodification of cognate trnas, leading to the exchange of g34 by preq1 at the wobble position in the anticodon loop. mutation of the tgt gene in shigella flexneri results in a significant loss of pathogenicity of the bacterium due to inefficient translation of a virulence protein mrna. herein, we describe the discovery of a ligand with an unexpected binding mode. on the basis of this binding mode, three slightly deviating ph ...200312646024
atmospheric pressure chemical ionisation reversed-phase liquid chromatography/ion trap mass spectrometry of intact bacteriohopanepolyols.atmospheric pressure chemical ionisation liquid chromatography/multi-stage ion trap mass spectrometry (apci-lc/ms(n)) has been applied to the study of intact bacteriohopanepolyols. spectral characterisation of bacteriohopanepolyols of known structure present in bacterial extracts (zymomonas mobilis and a fermenter containing methanotrophs including methylococcus capsulatus) has revealed greater structural detail than previous liquid chromatography/mass spectrometry (lc/ms) methods and identified ...200312661028
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
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