export of the periplasmic nadp-containing glucose-fructose oxidoreductase of zymomonas mobilis.glucose-fructose oxidoreductase (gfor) of the gram-negative bacterium zymomonas mobilis is a periplasmic enzyme with the tightly bound cofactor nadp. the preprotein carries an unusually long n-terminal signal sequence of 52 amino acid residues. a sorbitol-negative mutant strain (acm3963) was found to be deficient in gfor activity and was used for the expression of plasmid-borne copies of the wild-type gfo gene or of alleles encoding alterations in the signal sequence of the pre-gfor protein. z. ...19968661942
the relationship between growth enhancement and pet expression in escherichia coli.the pet operon consists of genes coding for enzymes responsible for ethanol production and consists of pyruvate dehydrogenase and alcohol dehydrogenase ii from the high-performance ethanologen zymomonas mobilis. this article describes the physiological influence of pet expression in escherichia coli b (atcc 11303) in terms of growth rate and overall concentrations of cell mass and catabolic end products achieved under well-defined cultivation conditions that included constant ph and carbon (ener ...19968669901
factors contributing to the loss of ethanologenicity of escherichia coli b recombinants pl0i297 and be economic and to be compatible with modern continuous bioconversion systems, it is imperative that the process organism exhibits an extremely high degree of stability. in the case of ethanol production from lignocellulosic biomass, functional stability of the potential process biocatalyst can be assessed in terms of the capacity to sustain high-performance fermentation during the continuous fermentation of biomass-derived sugars. this investigation employed glucose- or xylose-limited chemos ...19968669902
molecular characterization of the levansucrase gene from the endophytic sugarcane bacterium acetobacter diazotrophicus srt4.the acetobacter diazotrophicus srt4 gene encoding levansucrase (ec (isda) was isolated from a genomic library. the nucleotide sequence of a 2.3 kb dna fragment sufficient for complementation of a levansucrase-deficient mutant (obtained by ems treatment) was determined. the isda gene (1751 bp) coded for a polypeptide of molecular mass 64.9 kda with an isoelectric point of 5.2. the n-terminal amino acid sequence of the extracellular levansucrase indicated the presence of a precursor prot ...19968704949
6-phosphogluconate dehydratase from zymomonas mobilis: an iron-sulfur-manganese enzyme.the enzyme 6-phosphogluconate dehydratase has been isolated in a stable form by a simple one-step procedure using dye ligand chromatography. the role of metal ions in the activity and stability of the enzyme was investigated. as with aconitase and several other dehydratase enzymes, the active site includes an fe4s4 cluster. in addition, the purified enzyme has been shown to contain one manganese ion per subunit, which is also essential for activity. rapid inactivation by superoxide radical was o ...19968728108
identification and characterization of phon-sf, a gene on the large plasmid of shigella flexneri 2a encoding a nonspecific phosphatase.a gene encoding a nonspecific phosphatase, named phon-sf, was identified on the large virulence plasmid (pmysh6000) of shigella flexneri 2a ysh6000. the phosphatase activity in ysh6000 was observed under high-phosphate conditions. however, it was found that low-phosphate conditions induced a slightly higher level of activity. the nucleotide sequence of the phon-sf region cloned from pmysh6000 possessing the phon-sf gene encoded 249 amino acids with a typical signal sequence at the n terminus. th ...19968755883
ethanol production by a mixed culture of flocculent strains of zymomonas mobilis and saccharomyces sp.pure and mixed cultures of zymomonas mobilis and saccharomyces sp. were tested for the production of ethanol using sucrose as the carbon source. both strains, isolated from spontaneously fermenting sugar-cane juice, are flocculent and alcohol-tolerant. the best results were obtained using a mixed culture, with a yield of 0.5 g ethanol/g sugar consumed and a volumetric productivity of 1.5 g ethanol l-1 h-1. no levan was produced even if a sucrose-based medium was used.19968766695
overexpression, purification, and generation of a thermostable variant of zymomonas mobilis fructokinase.the gene encoding fructokinase (ec from zymomonas mobilis has been expressed at high level in escherichia coli by modifying the ribosome binding site using the polymerase chain reaction. a simple two-step purification from extracts of the recombinant cells results in highly purified enzyme suitable for use in fructose determination. using the polymerase chain reaction in mutagenic conditions, a variant of fructokinase was isolated which was more thermostable than the wild type, taking t ...19968776754
the genus sphingomonas: physiology and ecology.exploitation of the metabolic capabilities of the genus sphingomonas could provide important commercial benefits to biotechnology. recent advances have demonstrated that these organisms have unique abilities to degrade refractory contaminants, to serve as bacterial antagonists to phytopathogenic fungi, and to secrete the highly useful gellan exopolysaccharides. unfortunately, sphingomonas are also animal pathogens and can readily degrade the copper pipes in drinking water distribution systems. t ...19968785434
d-cycloserine biases enrichment for auxotrophic mutants of zymomonas mobilis.contrary to its effect on rich medium, d-cycloserine showed no bactericidal effect on zymomonas mobilis cells cultured on mineral medium. addition of a mixture of glycine and glutamic acid to the mineral medium restored its bactericidal action. however, mutant enrichments run in these conditions were biased, with mostly methionine mutants isolated. a decrease of the d-cycloserine concentration only reduced the bias.19968810500
cloning and expression of the zymomonas mobilis "production of ethanol" genes in lactobacillus casei.this study describes the expression of the zymomonas mobilis genes coding for pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adh) in lactobacillus casei 686. to promote transcription, the promoter and ribosome binding site (rbs) from the lactococcus lactis subsp. lactis-derived vector, pmge36e, were inserted upstream of the pdc gene. the former sequences were positioned such that translation of pdc was coupled to translation of an 81-base pair open reading frame terminating within the p ...19968824172
nucleotide sequence determination and genetic analysis of the bacteroides plasmid, pbi143.the nucleotide sequence and genetic organization of the bacteroides plasmid pbi143 were determined. the plasmid was 2747 base pairs (bp) and had a g+c content of 41% (genbank accession no. u30316). there were two open reading frames greater than 50 codons and these were designated moba and repa. a 56-bp inverted repeat divided pbi143 into modules with repa and moba in separate regions. there was a marked difference in the g+c content and codon usage for the two regions; repa had 33% g+c and moba ...19958825374
purification, crystallization, and preliminary x-ray diffraction studies of trna-guanine transglycosylase from zymomonas mobilis.the trna modifying enzyme trna-gnanine transglycosylase (tgt) catalyzes the exchange of guanine in the first position of the anticodon with the quenine precursor 7-aminomethyl-7-deazagnanine. tgt from zymomonas mobilis has been purified by crystallization and further recrystallized to obtain single crystals suitable for x-ray diffraction studies. crystals were grown by vapor diffusion/gel crystallization methods using peg 8,000 as precipitant. macroseeding techniques were employed to produce lar ...19968860000
expression of the escherichia coli pmi gene, encoding phosphomannose-isomerase in zymomonas mobilis, leads to utilization of mannose as a novel growth substrate, which can be used as a selective marker.wild-type zymomonas mobilis can utilize only three substrates (sucrose, glucose, and fructose) as sole carbon sources, which are largely converted into ethanol and carbon dioxide. here, we show that although d-mannose is not used as a growth substrate, it is taken up via the glucose uniport system (glucose facilitator protein) with a vmax similar to that of glucose. moreover, d-mannose was phosphorylated by a side activity of the resident fructokinase to mannose-6-phosphate. fructokinase was pur ...19968900006
specific spoilage organisms in breweries and laboratory media for their detection.the gram positive bacteria are generally regarded as the most hazardous beer spoilage organisms in modern breweries, especially the lactobacilli: l. brevis, l. lindneri, l. curvatus, l. casei, l. buchneri, l. coryneformis, l. plantarum, l. brevisimilis, l. malefermentans and l. parabuchneri and the pediococci: p damnosus, p. inopinatus and p. dextrinicus. micrococcus kristinae is the only species within the micrococci relevant to brewing. the gram negative strictly anaerobic bacteria are apparen ...19968913814
genetic and physiological analysis of the lethal effect of l-(+)-lactate dehydrogenase deficiency in streptococcus mutans: complementation by alcohol dehydrogenase from zymomonas mobilis.ch4ts is a previously isolated recombinant mutant of streptococcus mutans ng8 which produces a thermolabile l-(+)-lactate dehydrogenase (ldh) activity. it does not grow at 42 degrees c under a variety of cultivation conditions. in this study, we show that a batch culture of ch4ts shifted from 30 to 42 degrees c underwent rapid cessation of growth and accelerated cell death. the mutant grew at 42 degrees c in continuous culture under glucose-limiting conditions. under these conditions, lactate pr ...19968926105
development of an arabinose-fermenting zymomonas mobilis strain by metabolic pathway engineering.the substrate fermentation range of the ethanologenic bacterium zymomonas mobilis was expanded to include the pentose sugar, l-arabinose, which is commonly found in agricultural residues and other lignocellulosic biomass. five genes, encoding l-arabinose isomerase (araa), l-ribulokinase (arab), l-ribulose-5-phosphate-4-epimerase (arad), transaldolase (talb), and transketolase (tkta), were isolated from escherichia coli and introduced into z. mobilis under the control of constitutive promoters th ...19968953718
stabilization of pet operon plasmids and ethanol production in escherichia coli strains lacking lactate dehydrogenase and pyruvate formate-lyase the last decade, a major goal of research in biofuels has been to metabolically engineer microorganisms to ferment multiple sugars from biomass or agricultural wastes to fuel ethanol. escherichia coli strains genetically engineered to contain the pet operon (zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase b genes) produce high levels of ethanol. strains carrying the pet operon in plasmid (e.g., e. coli b/ploi297) or in chromosomal (e.g., e. coli ko11) sites require antibiot ...19968953729
production of levan, a fructose polymer, using an overexpressed recombinant levansucrase. 19968958116
mutagenesis and crystallographic studies of zymomonas mobilis trna-guanine transglycosylase reveal aspartate 102 as the active site nucleophile.procaryotic trna-guanine transglycosylase (tgt) catalyzes the posttranscriptional base exchange of the queuine precursor 7-aminomethyl-7-deazaguanine (preq1) with the genetically encoded guanine at the wobble position of trnas specific for asn, asp, his, and tyr. the x-ray structures of zymomonas mobilis tgt and of its complex with preq1 [romier, c., reuter, k., suck, d., & ficner, r. (1996) embo j. 15, 2850-2857] have revealed a specific preq1 binding pocket and allowed a proposal for trna bind ...19968961936
the structure of glucose-fructose oxidoreductase from zymomonas mobilis: an osmoprotective periplasmic enzyme containing non-dissociable nadp.the organism zymomonas mobilis occurs naturally in sugar-rich environments. to protect the bacterium against osmotic shock, the periplasmic enzyme glucose-fructose oxidoreductase (gfor) produces the compatible, solute sorbitol by reduction of fructose, coupled with the oxidation of glucose to gluconolactone. hence, z mobilis can tolerate high concentrations of sugars and this property may be useful in the development of an efficient microbial process for ethanol production. each enzyme subunit c ...19968994968
differential inactivation of alcohol dehydrogenase isoenzymes in zymomonas mobilis by oxygen.zymomonas mobilis is endowed with two isoenzymes of fermentative alcohol dehydrogenase, a zinc-containing enzyme (adh i) and an iron-containing enzyme (adh ii). the activity of adh i remains fully conserved, while adh ii activity decays when anaerobic cultures are shifted to aerobiosis. this differential response depends on the metal present on each isoenzyme, since pure preparations of adh i are resistant to oxidative inactivation and preparations of zinc-containing adh ii, obtained by incubati ...19979023190
an elevation of the molar growth yield of zymomonas mobilis during aerobic exponential growthelevated values of molar growth yield (yx/s = 14-26 g mol-1) were obtained during exponential growth (μ > 0.4 h-1) of zymomonas mobilis atcc 29191 by using reduced concentrations of glucose (6.25-100 mm) and increased oxygen supply (eh > 300 mv) in the growth medium, as compared to the yx/s of anaerobic exponential growth (8-10 g mol-1). aerobically grown cells showed an increased maximum growth rate (μmax), and a reduced specific glucose consumption rate (qs), and specific ethanol formation rat ...19979042757
cysteine 265 is in the active site of, but is not essential for catalysis by trna-guanine transglycosylase (tgt) from escherichia mutagenesis and x-ray absorption spectroscopy studies have previously shown that the trna-guanine transglycosylase (tgt) from escherichia coli is a zinc metalloprotein and identified the enzymic ligands to the zinc [chong et al. (1995), biochemistry 34, 3694-3701; garcia et al. (1966), biochemistry 35, 3133-3139]. during these studies one mutant, tgt (c265a), was found to exhibit a significantly lower specific activity, but was not found to be involved in the zinc site. the present ...19979055203
nicotinoprotein alcohol/aldehyde oxidoreductases. enzymes with bound nad(p) as cofactor. 19979059647
expression of the zymomonas mobilis gfo gene or nadp-containing glucose:fructose oxidoreductase (gfor) in escherichia coli. formation of enzymatically active pregfor but lack of processing into a stable periplasmic protein.glucose:fructose oxidoreductase (gfor) of the gram-negative bacterium zymomonas mobilis is a periplasmic enzyme with tightly bound cofactor nadp. the preprotein carries an unusually long n-terminal signal peptide of 52 amino acid residues. expression of the gfo gene in cells of escherichia coli k12, under the control of a tac promoter, led to immunologically detectable proteins in western blots, and to the formation of an enzymatically active precursor form (pregfor), located in the cytosol. pro ...19979063452
extracellular melibiose and fructose are intermediates in raffinose catabolism during fermentation to ethanol by engineered enteric bacteria.contrary to general concepts of bacterial saccharide metabolism, melibiose (25 to 32 g/liter) and fructose (5 to 14 g/liter) accumulated as extracellular intermediates during the catabolism of raffinose (o-alpha-d-galactopyranosyl-1, 6-alpha-d-glucopyranosyl-beta-d-fructofuranoside) (90 g/liter) by ethanologenic recombinants of escherichia coli b, klebsiella oxytoca m5a1, and erwinia chrysanthemi ec16. both hydrolysis products (melibiose and fructose) were subsequently transported and further me ...19979068632
sampling tube device for monitoring intracellular metabolite dynamics.continuous sampling of microorganisms from a controlled bioreactor with rapid inactivation of metabolism and extraction of metabolites using precooled -40 degrees c perchloric acid solution (35%) was achieved with a sampling tube, thus fixing fast dynamic reactions at a certain position in the tube. after sampling was stopped (200 s) the tube was frozen at -80 degrees c and divided into identical parts and the extracted metabolites were analyzed enzymatically. a high resolution in time was achie ...19979073360
protein design on pyruvate decarboxylase (pdc) by site-directed mutagenesis. application to mechanistical investigations, and tailoring pdc for the use in organic synthesis.pyruvate decarboxylases (e.c. from various organisms have been studied for many years, mainly with respect to the mechanism of the non-oxidative decarboxylation reaction. although the c-c-bond-forming properties of these enzymes are known and have been applied for many years in biotransformations for the synthesis of chiral alpha-hydroxy ketones, only little is known about the factors influencing the carboligase side-reaction. the present review surveys recent efforts in the study of si ...19979103910
alteration of substrate specificity of zymomonas mobilis alcohol dehydrogenase-2 using in vitro random mutagenesis.random mutagenesis of the gene encoding zymomonas mobilis alcohol dehydrogenase-2 has enabled isolation of variants of the enzyme that have substrate specificities different from that of the wild-type enzyme. after amino acids responsible for the changes were identified, directed mutation at these sites was also carried out. variants that are active on butanol have been investigated in detail. changes at residue 161 and other changes at residues 155 and 165 cause enhanced activity with longer-ch ...19979116506
an elevation of the molar growth yield of zymomonas mobilis during aerobic exponential growth.elevated values of molar growth yield (yx/s = 14-26 g mol-1) were obtained during exponential growth (mu > 0.4 h-1) of zymomonas mobilis atcc 29191 by using reduced concentrations of glucose (6. 25-100 mm) and increased oxygen supply (eh > 300 mv) in the growth medium, as compared to the yx/s of anaerobic exponential growth (8-10 g mol-1). aerobically grown cells showed an increased maximum growth rate (mumax), and a reduced specific glucose consumption rate (qs), and specific ethanol formation ...19979133324
modulation of lipoxygenase activity by bacterial hopanoids.tetrahydroxybacteriohopane (1), a bacterial hopanoid, inhibited soybean 15-lipoxygenase with an ic50 of about 10 microm. after per-o-acetylation of 1 no inhibition of the 15-lipoxygenase was observed. two other bacterial hopanoids, tetrahydroxybacteriohopane glucosamine (2) and tetrahydroxybacteriohopane ether (3), stimulated the activity of soybean 15-lipoxygenase. the activities of two other arachidonic acid-metabolizing enzymes, human 5-lipoxygenase and prostaglandin h synthase, were unaffect ...19979134748
a glutamate uptake regulatory protein (grp) in escherichia coli? 19979140981
squalene-hopene cyclase from bradyrhizobium japonicum: cloning, expression, sequence analysis and comparison to other triterpenoid cyclases.with the help of a pcr-based screening method, the gene encoding squalenehopene cyclase (shc) of bradyrhizobium japonicum usda 110 was isolated from a cosmid library. the shc catalyses the cyclization of squalene to hopanoids, a class of triterpenoid lipids recently discovered in nitrogen-fixing, root-nodule-forming bradyrhizobium bacteria. hybridization experiments showed that the gene is present in bacteria of all bradyrhizobium strains tested and in photosynthetic bacteria forming stem nodule ...19979141686
the substitution of a single amino acid residue (ser-116 --> asp) alters nadp-containing glucose-fructose oxidoreductase of zymomonas mobilis into a glucose dehydrogenase with dual coenzyme specificity.glucose-fructose oxidoreductase (gfor, ec from the gram-negative bacterium zymomonas mobilis contains the tightly bound cofactor nadp. based on the revision of the gfo dna sequence, the derived gfor sequence was aligned with enzymes catalyzing reactions with similar substrates. a novel consensus motif (agkhvxcekp) for a class of dehydrogenases was detected. from secondary structure analysis the serine-116 residue of gfor was predicted as part of a rossmann-type dinucleotide binding f ...19979148926
antimicrobial activity of flavonoids from leaves of tagetes minuta.the total extract and fractions with different solvents, obtained from leaves of tagetes minuta, showed several degrees of antimicrobial activity against gram positive and gram negative microorganisms. the same fractions were inactive against lactobacillus, zymomonas and saccharomices species. the major component of the extract: quercetagetin-7-arabinosyl-galactoside, showed significant antimicrobial activity on pathogen microorganisms tested. correlation results were carried out using chloramph ...19979201613
changes in the growth and enzyme level of zymomonas mobilis under oxygen-limited conditions at low glucose concentration.zymomonas mobilis growing aerobically with 20 g glucose-1 (carbon-limited) in a chemostat exhibited an increase in both the molar growth yield (yx/s) and the maximum molar growth yield (yx/smax) and a decrease in both the specific substrate consumption rate (qs) and the maintenance energy consumption rate (me). stepwise increase in the input oxygen partial pressure showed that anaerobic-to-aerobic transitional adaptation occurred in four stages: anaerobic (0 mm hgo2), oxygen-limited (7.6- 230 mm ...19979211713
the effect of ethanol and oxygen on the growth of zymomonas mobilis and the levels of hopanoids and other membrane lipids.zymomonas mobilis (atcc 29191) was grown either aerobically or anaerobically in the presence of 2% (wt/vol) glucose and 0, 3, or 6% (vol/vol) ethanol. the rates of growth and the composition of hopanoids, cellular fatty acids, and other lipids in the bacterial membranes were quantitatively analyzed. the bacterium grew in the presence of 3% and 6% ethanol and was more ethanol tolerant when grown anaerobically. in the absence of ethanol, hopanoids comprised about 30% (by mass) of the total cellula ...19979216888
allosteric control of zymomonas mobilis glucose-6-phosphate dehydrogenase by phosphoenolpyruvate.the second enzyme of the entner-doudoroff glycolytic pathway in zymomonas mobilis, glucose-6-phosphate dehydrogenase, has been found to be inhibited by phosphoenolpyruvate (pep). in the presence of pep levels in the micromolar range, the response of the enzyme to glucose 6-phosphate concentration becomes sigmoidal, with a hill coefficient up to 2. at low ionic strength in the absence of pep, the response to glucose 6-phosphate concentration is michaelis-menten, but at physiological ionic strengt ...19979307022
the role of his113 and his114 in pyruvate decarboxylase from zymomonas mobilis.pyruvate decarboxylase (pdc) is one of several enzymes that require thiamin diphosphate (thdp) and a divalent cation as essential cofactors. recently, the three-dimensional structures of the enzyme from two yeasts have been determined. while these structures shed light on the binding of the cofactors and the reaction mechanism, the interactions between the substrate pyruvate and the enzyme remain unclear. we have used pdc from zymomonas mobilis as a model for these enzymes in order to study subs ...19979310361
the cis-diol dehydrogenase cbac gene of tn5271 is required for growth on 3-chlorobenzoate but not 3,4-dichlorobenzoate.the nucleotide sequence of cbac, the 1-carboxy-3-chloro-4,5-dihydroxycyclohexa-2,6-diene (cis-diol) dehydrogenase gene from the 3-chlorobenzoate (3-cba) catabolic transposon tn5271 was determined. the functional significance of the deduced open reading frame was evaluated by deletion of an internal bsteii restriction site in cbac and by the creation of nested deletions using exonuclease iii. expression studies were carried out with alcaligenes sp. strain br6024, a chloramphenicol-resistant, tryp ...19979322760
isolation and molecular characterization of high-performance cellobiose-fermenting spontaneous mutants of ethanologenic escherichia coli ko11 containing the klebsiella oxytoca casab operon.escherichia coli ko11 was previously constructed to produce ethanol from acid hydrolysates of hemicellulose (pentoses and hexoses) by the chromosomal integration of zymomonas mobilis genes encoding pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adhb). klebsiella oxytoca p2 was constructed in an analogous fashion for the simultaneous saccharification and fermentation of cellulose and contains pts enzymes for cellobiose. in this study, ko11 was further engineered for the fermentation of c ...19979406380
measurement of platelet aggregation peptide inhibitors by ultrasonic interferometry.several peptide inhibitors of thrombin- or collagen-induced platelet aggregation and of the interaction between glycoprotein ib and von willebrand factor were studied by a new method--ultrasonic interferometry (echo cell). inhibition of aggregate formation in a concentration-dependent manner was observed. the sensitivity of the method was 3 to 40 times higher than that of classical turbidimetry.19989451507
a physical map of the genome of ethanol fermentative bacterium zymomonas mobilis zm4 and localization of genes on the map.a physical map of the zymomonas mobilis zm4 genome has been constructed from the results of reciprocal southern hybridization with pmei, paci, and noti-digested genomic dna fragments and linking cosmid clones. restriction enzyme-digested z. mobilis zm4 genome was electrophoresed with phage lambda dna concatemers as a size standard in a bio-rad chef-drii pulsed-field gel electrophoresis (pfge) system. the restriction enzyme pmei generated 15 fragments (3-625 kb), and paci produced 19 fragments (7 ...19989469936
thermostable variants of zymomonas mobilis alcohol dehydrogenase obtained using pcr-mediated random mutagenesis.using a random mutagenesis technique, the ferrous-ion-activated alcohol dehydrogenase of zymomonas mobilis has been altered to produce more thermally stable variants. after three rounds of mutation, a variant over 10 degrees c more stable at ph 8, with essentially unaltered kinetic characteristics, was produced. however, the ph profile of thermostability of this variant was much altered compared with the wild-type, with a relatively small increase (4 degrees c) at ph 6. sequencing of the variant ...19989473458
a multistep process is responsible for product-induced inactivation of glucose-fructose oxidoreductase from zymomonas mobilis.glucose-fructose oxidoreductase from the bacterium zymomonas mobilis catalyzes a transhydrogenation reaction in which d-fructose reduction to d-sorbitol is coupled to the oxidation of d-glucose or other aldoses to the corresponding aldonolactones. tightly protein-bound nadp(h) serves as the cofactor. we found that the interaction of glucose-fructose oxidoreductase with its aldonolactone product triggered a sequential process that affects the protein structure conformationally and chemically and, ...19989490072
the effects of temperature on the kinetics and stability of mesophilic and thermophilic 3-phosphoglycerate kinases.the effects of temperature on the kinetic parameters kcat and km, for three isolates of the highly conserved monomeric enzyme 3-phosphoglycerate kinase (pgk), were investigated in detail using a rapid automated kinetics apparatus. pgk was purified from the thermophilic bacterium thermoanaerobacter sp. rt8.g4 (optimum growth temperature 68 degrees c), the mesophile zymomonas mobilis (optimum growth temperature 32 degrees c) and a second, unidentified, soil mesophile designated unid a (optimum gro ...19989494072
purification of the pyruvate dehydrogenase multienzyme complex of zymomonas mobilis and identification and sequence analysis of the corresponding genes.the pyruvate dehydrogenase (pdh) complex of the gram-negative bacterium zymomonas mobilis was purified to homogeneity. from 250 g of cells, we isolated 1 mg of pdh complex with a specific activity of 12.6 u/mg of protein. analysis of subunit composition revealed a pdh (e1) consisting of the two subunits e1alpha (38 kda) and e1beta (56 kda), a dihydrolipoamide acetyltransferase (e2) of 48 kda, and a lipoamide dehydrogenase (e3) of 50 kda. the e2 core of the complex is arranged to form a pentagona ...19989515924
metabolic engineering for the production of carotenoids in non-carotenogenic bacteria and yeasts.the crt gene clusters responsible for the biosynthesis of carotenoids such as lycopene, beta-carotene and astaxanthin have been isolated from carotenogenic bacteria such as erwinia species and the marine bacterium agrobacterium aurantiacum. the functions of the individual genes have been identified. the first substrate of the enzymes encoded by the erwinia crt clusters is farnesyl pyrophosphate which is not only the precursor for carotenoid biosynthesis but also sterols, dolichols and other nume ...19979519479
small-angle x-ray solution-scattering studies on ligand-induced subunit interactions of the thiamine diphosphate dependent enzyme pyruvate decarboxylase from different organisms.the quaternary structures of the thiamine diphosphate dependent enzyme pyruvate decarboxylase (ec from the recombinant wild type of saccharomycescerevisiae and zymomonas mobilis and from germinating pisum sativum seeds were examined by x-ray solution scattering. the dependence of the subunit association equilibrium on the ph and the presence of the cofactors thiamine diphosphate and magnesium ions were compared, and the differences between the catalytic properties of the different enzym ...19989548765
purification and characterization of a novel levanoctaose-producing levanase from pseudomonas strain k-52.levan-assimilating micro-organisms from soil samples were screened for levanoligosaccharide-generating enzyme production. the isolated strain k-52 produced an extracellular levanoctaose-generating enzyme and was identified as belonging to genus pseudomonas. the levanase was purified to homogeneity by (nh4)2so4 fractionation and successive column chromatography on deae-cellulose, phenyl-toyopearl 650 m, sephadex g-100 and hydroxyapatite. the molecular mass of the enzyme was estimated as approx. 3 ...19989569612
the dna-binding protein ii from zymomonas mobilis. complete amino acid sequence and interaction with dna.the primary structure of the dna-binding protein ii from zymomonas mobilis has been determined from data provided by automated edman degradation of the intact protein and of peptides derived from cleavage at aspartic acid and arginine residues. when compared with the homologous protein isolated from other bacteria, the dna-binding protein ii from z mobilis shows many substitutions. several non-conservative substitutions at positions usually highly conserved in this type of protein probably accou ...19989587668
isolation and characterization of ethanol-tolerant mutants of escherichia coli ko11 for fuel ethanol production.genetically engineered escherichia coli ko11 is capable of efficiently producing ethanol from all sugar constituents of lignocellulose but lacks the high ethanol tolerance of yeasts currently used for commercial starch-based ethanol processes. using an enrichment method which selects alternatively for ethanol tolerance during growth in broth and for ethanol production on solid medium, mutants of ko11 with increased ethanol tolerance were isolated which can produce more than 60 g ethanol l-1 from ...19989611822
surface display of zymomonas mobilis levansucrase by using the ice-nucleation protein of pseudomonas syringae.the ice-nucleation protein (inp) is a glycosyl phosphatidylinositol-anchored outer membrane protein found in some gram-negative bacteria. using pseudomonas syringae inp as an anchoring motif, we investigated the functional display of a foreign protein, zymomonas mobilis levansucrase (levu), on the surface of escherichia coli. the cells expressing inp-levu were found to retain both the ice-nucleation and whole-cell levansucrase enzyme activities, indicating the functional expression of inp-levu h ...19989624691
improving fermentation performance of recombinant zymomonas in acetic acid-containing the production of ethanol from lignocellulosic biomass, the hydrolysis of the acetylated pentosans in hemicellulose during pretreatment produces acetic acid in the prehydrolysate. the national renewable energy laboratory (nrel) is currently investigating a simultaneous saccharification and cofermentation (sscf) process that uses a proprietary metabolically engineered strain of zymomonas mobilis that can coferment glucose and xylose. acetic acid toxicity represents a major limitation to biocon ...19989627380
conditions that promote production of lactic acid by zymomonas mobilis in batch and continuous culture.this study documents the similar ph-dependent shift in pyruvate metabolism exhibited by zymomonas mobilis atcc 29191 and atcc 39676 in response to controlled changes in their steady-state growth environments. the usual high degree of ethanol selectivity associated with glucose fermentation by z. mobilis is associated with conditions that promote rapid and robust growth, with about 95% of the substrate (5% w/v glucose) being converted to ethanol and c)2, and the remaining 5% being used for the sy ...19989627381
cloning of a sphingomonas paucimobilis syk-6 gene encoding a novel oxygenase that cleaves lignin-related biphenyl and characterization of the enzyme.sphingomonas paucimobilis syk-6 transforms 2,2'-dihydroxy-3,3'-dimethoxy-5,5'-dicarboxybiphenyl (ddva), a lignin-related biphenyl compound, to 5-carboxyvanillic acid via 2,2',3-trihydroxy-3'-methoxy-5,5'-dicarboxybiphenyl (oh-ddva) as an intermediate (15). the ring fission of oh-ddva is an essential step in the ddva degradative pathway. a 15-kb ecori fragment isolated from the cosmid library complemented the growth deficiency of a mutant on oh-ddva. subcloning and deletion analysis showed that a ...19989647824
increased cellulose production from sucrose with reduced levan accumulation by an acetobacter strain harboring a recombinant plasmid.cellulose production from sucrose by acetobacter strains is accompanied by the accumulation of a water-soluble polysaccharide, called levan. to improve cellulose productivity, a levansucrase-deficient mutant, ld-2, was derived from acetobacter strain 757 and used as a host for the construction of recombinant strains. an ld-2 mutant harboring a plasmid containing the sucrase gene, sucze3, from zymomonas mobilis together with zlis, a gene that encodes a secretion-activating factor under the contro ...19989648211
purification and characterization of alkaline phosphatase containing phosphotyrosyl phosphatase activity from the bacterium prevotella intermedia.a novel alkaline phosphatase, designated pialp, has been purified and characterized from prevotella intermedia atcc 25611, an anaerobe implicated in progressive periodontal disease. the enzyme was a homodimer of apparently identical subunits of mr 54 kda. thiol-reducing agents completely inhibited the purified enzyme. the enzyme was highly stable even at 80 degrees c. it exhibited substantial activity against tyrosine-phosphate-containing raytide. the phosphatase activity was sensitive to orthov ...19989654126
transition-state theoretical interpretation of the catalytic power of pyruvate decarboxylases: the roles of static and dynamical considerations.the catalytic power of two thiamin diphosphate (thdp)-dependent enzymes, yeast pyruvate decarboxylase (the hysteretically regulated enzyme from saccharomyces cerevisiae, scpdc) and bacterial pyruvate decarboxylase (the unregulated enzyme from zymomonas mobilis, zmpdc), are analyzed by thorough-going application of transition-state theory, i.e. by a static approach that emphasizes the state-function character of the free energy of activation and takes no explicit account of dynamical consideratio ...19989655907
subunit structure, function and organisation of pyruvate decarboxylases from various organisms.the nature of the environment of macromolecules influences and determines the state of their overall structure and the extent of binding of specific (cofactors, substrates) or unspecific ligands. how these interactions between enzyme molecules and ligands influence their quaternary structures and, in this way, the realisation of high catalytic activity will be discussed here for the enzyme pyruvate decarboxylase from various organisms: brewer's yeast, brewer's yeast strain, recombinant wild type ...19989655918
structure and properties of pyruvate decarboxylase and site-directed mutagenesis of the zymomonas mobilis enzyme.pyruvate decarboxylase (ec is a thiamin diphosphate-dependent enzyme that catalyzes the penultimate step in alcohol fermentation. the enzyme is widely distributed in plants and fungi but is rare in prokaryotes and absent in animals. here we review its structure and properties with particular emphasis on how site-directed mutagenesis of the enzyme from zymomonas mobilis has assisted us to understand the function of critical residues.19989655927
gene and subunit organization of bacterial pyruvate dehydrogenase complexes.pyruvate dehydrogenase complexes of bacterial origin are compared with respect to subunit composition, organization of the corresponding genes, and the number and location of lipoyl domains. special attention is given to two unusual examples of pyruvate dehydrogenase complexes, formed by zymomonas mobilis and thiobacillus ferrooxidans.19989655937
high resolution crystal structure of pyruvate decarboxylase from zymomonas mobilis. implications for substrate activation in pyruvate decarboxylases.the crystal structure of tetrameric pyruvate decarboxylase from zymomonas mobilis has been determined at 1.9 a resolution and refined to a crystallographic r-factor of 16.2% and rfree of 19.7%. the subunit consists of three domains, all of the alpha/beta type. two of the subunits form a tight dimer with an extensive interface area. the thiamin diphosphate binding site is located at the subunit-subunit interface, and the cofactor, bound in the v conformation, interacts with residues from the n-te ...19989685367
control of the association state of tetrameric glucose-fructose oxidoreductase from zymomonas mobilis as the rationale for stabilization of the enzyme in biochemical reactors.tetrameric, nadp-containing glucose-fructose oxidoreductase (gfor) from zymomonas mobilis catalyzes the oxidation of glucose into glucono-delta-lactone coupled to the reduction of fructose to sorbitol. gfor is inactivated during substrate turnover in vitro, the long-term stability of the enzyme during conversions in biochemical reactors thereby being drastically reduced. the process of inactivation is triggered by structural transitions that are induced by the lactone product and involves aggreg ...19989685715
cloning of conserved genes from zymomonas mobilis and bradyrhizobium japonicum that function in the biosynthesis of hopanoid lipids.the squalene-hopene cyclase (shc) is the only enzyme involved in the biosynthesis of hopanoid lipids that has been characterized on the genetic level. to investigate if additional genes involved in hopanoid biosynthesis are clustered with the shc gene, we cloned and analyzed the nucleotide sequences located immediately upstream of the shc genes from zymomonas mobilis and bradyrhizobium japonicum. in z. mobilis, five open reading frames (orfs, designated as hpna-e) were detected in a close arrang ...19989714766
cloning, nucleotide sequence, and expression in escherichia coli of levansucrase genes from the plant pathogens pseudomonas syringae pv. glycinea and p. syringae pv. phaseolicola.plant-pathogenic bacteria produce various extracellular polysaccharides (epss) which may function as virulence factors in diseases caused by these bacteria. the eps levan is synthesized by the extracellular enzyme levansucrase in pseudomonas syringae, erwinia amylovora, and other bacterial species. the lsc genes encoding levansucrase from p. syringae pv. glycinea pg4180 and p. syringae pv. phaseolicola ncppb 1321 were cloned, and their nucleotide sequences were determined. heterologous expressio ...19989726857
no prokaryotic gpi anchoring. 19989743095
cloning and expression of the zymomonas mobilis pyruvate kinase gene in escherichia coli.the homotetrameric pyruvate kinases (pk) constitute a fine example of allosteric enzymes subjected to sophisticated regulatory mechanisms. we have cloned and sequenced the zymomonas mobilis structural gene for the first prokaryotic dimeric pk, as an initial step toward understanding the peculiar properties of this enzyme. the deduced amino acid sequence of the pyk gene consists of 475 residues with a calculated molecular mass of 51.4kda and exhibits up to 50% sequence identity with other pks. he ...19989767092
novel selection for isoniazid (inh) resistance genes supports a role for nad+-binding proteins in mycobacterial inh resistance.the genetic basis of isoniazid (inh) resistance remains unknown for a significant proportion of clinical isolates. to identify genes which might confer resistance by detoxifying or sequestering inh, we transformed the escherichia coli oxyr mutant, which is relatively sensitive to inh, with a mycobacterium tuberculosis plasmid library and selected for inh-resistant clones. three genes were identified and called ceo for their ability to complement the escherichia coli oxyr mutant. ceoa was the pre ...19989784509
lactone-ring-cleaving enzyme: genetic analysis, novel rna editing, and evolutionary implications.a lactonohydrolase from fusarium oxysporum aku 3702 is an enzyme catalyzing the hydrolysis of aldonate lactones to the corresponding aldonic acids. the amino acid sequences of the nh2 terminus and internal peptide fragments of the enzyme were determined to prepare synthetic oligonucleotides as primers for the pcr. an approximate 1, 000-base genomic dna fragment thus amplified was used as the probe to clone both genomic dna and cdna for the enzyme. the lactonohydrolase genomic gene consists of si ...19989788992
expression of the extracellular levansucrase and invertase genes from zymomonas mobilis in escherichia coli cells.we investigated the expression and localization in escherichia coli of sucze2 and sucze3, encoding zymomonas mobilis extracellular levansucrase and invertase, respectively, and lacking a typical n-terminal secretion signal. levansucrase and invertase were expressed efficiently under the lac and tac promoters in e. coli cells, and some of the levansucrase produced was localized in the periplasmic space. the sucze2 expression was not lethal to e. coli in the presence of 5% sucrose, and led to the ...19989805385
membrane d-lactate oxidase in zymomonas mobilis: evidence for a branched respiratory chain.respiratory chain composition of the ethanol-producing bacterium zymomonas mobilis was studied. its membrane d-lactate oxidase was characterised. with nadh, but not d-lactate as substrate, a cytochrome o-like component was seen in co difference spectra. chlorpromazine specifically inhibited reduction of cytochrome d, while myxothiazol eliminated the cytochrome o-like features in co difference spectra. it is suggested that electrons from nadh are distributed between branches terminated by the cyt ...19989812368
active site mutants of pyruvate decarboxylase from zymomonas mobilis--a site-directed mutagenesis study of l112, i472, i476, e473, and n482.the homotetrameric pyruvate decarboxylase (pdc) from zymomonas mobilis requires the cofactors thiamin diphosphate and mg2+ for catalytic activity. we have investigated the role of various amino acid residues in the direct environment of the active site. the role of residue e473 in the catalytic activity and stability of the enzyme was probed by several mutations. all mutant enzymes were either inactive or failed to give any recombinant protein. the close interaction of e473 and n482, which can b ...19989839941
characterization of the mobilization region of the zymomonas mobilis atcc10988 plasmid pzmo3.the 2.7-kb zymomonas mobilis atcc10988 plasmid pzmo3 contains a coding region (orf1) indispensable for mobilization. a cis-acting 409-bp sequence between orf2 (c-terminal) and orf1 (n-terminal) conferred mobilization activity to puc19, when the product of orf1 was provided in trans. in this area, two segments showed homology with previously characterized orit regions.19999887309
activation of thiamine diphosphate in pyruvate decarboxylase from zymomonas mobilis.replacement of tryptophan 392 located in the active site cavity of pyruvate decarboxylase (pdc; ec from zymomonas mobilis by methionine or glutamine yields enzymes with smaller catalytic constants of 8.5 s(-1) and 3.6 s(-1) at 4 degrees c, compared to that of the wild-type enzyme (17 s(-1)). the rate constants of the h/d exchange at the c2 of the coenzyme thiamine diphosphate have been determined to be 130 s(-1) for the wild-type enzyme, 56 s(-1) for the methionine and 30 s(-1) for the ...19989891980
sequence analysis of a cryptic plasmid from flavobacterium sp. kp1, a psychrophilic bacterium.a cryptic plasmid found at high copy number was isolated from flavobacterium sp. kp1, a psychrophilic gram-negative bacterium, cloned, and sequenced. the sequence will appear in the ddbj/embl/genbank databases under the accession number ab007196. the pfl1 plasmid is 2311 nucleotides in length with 32.7% gc content, and shows a distinctive nucleotide sequence without homology to other plasmids of similar length. the plasmid contains two open reading frames of significant length, orfi and orfii. o ...19999919674
ethanol synthesis by genetic engineering in cyanobacteria.cyanobacteria are autotrophic prokaryotes which carry out oxygenic photosynthesis and accumulate glycogen as the major form of stored carbon. in this research, we introduced new genes into a cyanobacterium in order to create a novel pathway for fixed carbon utilization which results in the synthesis of ethanol. the coding sequences of pyruvate decarboxylase (pdc) and alcohol dehydrogenase ii (adh) from the bacterium zymomonas mobilis were cloned into the shuttle vector pcb4 and then used to tran ...19999925577
levansucrase of rahnella aquatilis atcc33071. gene cloning, expression, and levan formation. 19989928133
complement activation by bacterial surface glycolipids: a study with planar bilayer membranes.planar asymmetric glycolipid/phospholipid bilayer membranes were used as a reconstitution model of the lipid matrix of the outer membrane of gram-negative bacteria to study complement (c) activation by various bacterial surface glycolipids with the aim of defining the c activation pathway. as glycolipids the lipopolysaccharides of salmonella enterica serovar minnesota r mutant strains r595 (re lps) and r4 (rd2 lps), pentaacyl lipid a from the lps of the escherichia coli re mutant f515, and glyco ...19999929374
aspartate-27 and glutamate-473 are involved in catalysis by zymomonas mobilis pyruvate decarboxylase.zymomonas mobilis pyruvate decarboxylase (ec was subjected to site-directed mutagenesis at two acidic residues near the thiamin diphosphate cofactor in the active site. asp-27 was changed to glu or asn, and glu-473 was mutated to asp (e473d) or gln (e473q). each mutant protein was purified to near-homogeneity, and the kinetic and cofactor-binding properties were compared with those of the wild-type protein. despite the very conservative nature of these alterations, all mutants had a ver ...199910191255
application of mathematical tools for metabolic design of microbial ethanol production.many attempts to engineer cellular metabolism have failed due to the complexity of cellular functions. mathematical and computational methods are needed that can organize the available experimental information, and provide insight and guidance for successful metabolic engineering. two such methods are reviewed here. both methods employ a (log)linear kinetic model of metabolism that is constructed based on enzyme kinetics characteristics. the first method allows the description of the dynamic res ...199810191385
metabolic engineering of bacteria for ethanol productiontechnologies are available which will allow the conversion of lignocellulose into fuel ethanol using genetically engineered bacteria. assembling these into a cost-effective process remains a challenge. our work has focused primarily on the genetic engineering of enteric bacteria using a portable ethanol production pathway. genes encoding zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase have been integrated into the chromosome of escherichia coli b to produce strain ko11 for the ...199810191391
effect of calcium on the surfactant tolerance of a fluoranthene degrading bacterium.surfactants are known to increase the apparent aqueous solubility of polycyclic aromatic hydrocarbons (pahs) and may thus be used to enhance the bioavailability and thereby to stimulate the biodegradation of these hydrophobic compounds. however, surfactants may in some cases reduce or inhibit biodegradation because of toxicity to the bacteria. in this study, toxicity of surfactants on sphingomonas paucimobilis strain epa505 and the effect on fluoranthene mineralization were investigated using tr ...199810192897
cloning and sequencing of a protein involved in phagosomal membrane fusion in mab was raised to the c5 phagosomal antigen in paramecium multimicronucleatum. to determine its function, the cdna and genomic dna encoding c5 were cloned. this antigen consisted of 315 amino acid residues with a predicted molecular weight of 36,594, a value similar to that determined by sds-page. sequence comparisons uncovered a low but significant homology with a schizosaccharomyces pombe protein and the c-terminal half of the beta-fructofuranosidase protein of zymomonas mobilis. lacking an ...199910198055
in-vitro study of interaction between photooxidation and biodegradation of 2-methylphenanthrene by sphingomonas sp. 2mpii.this work reports a study of interactions between reactions of photooxidation and reactions of bacterial degradation of an alkylated polyaromatic hydrocarbon (2-methylphenanthrene). bacterial growth was carried out using artificial sunlight as light source. among the various products detected, the major product was identified as the 2-methylphenanthrene aldehyde. sunlight allows accelerated elimination of the substrate. this enhancement of the biodegradation rate of 2 methylphenanthrene is due t ...199910204235
bacterial proteins carrying twin-r signal peptides are specifically targeted by the delta ph-dependent transport machinery of the thylakoid membrane system.glucose-fructose oxidoreductase (gfor), a periplasmic protein of zymomonas mobilis, is synthesized as a precursor polypeptide with a twin-r signal peptide for sec-independent protein export in bacteria. in higher plant chloroplasts, twin-r signal peptides are specific targeting signals for the sec-independent delta ph pathway of the thylakoid membrane system. in agreement with the assumed common phylogenetic origin of the two protein transport mechanisms, gfor can be efficiently translocated by ...199910218590
transcriptional analysis of levu operon encoding saccharolytic enzymes and two apparent genes involved in amino acid biosynthesis in zymomonas mobilis.extracellular levansucrase (levu) and sucrase (invb) are two of the three saccharolytic enzymes involved in the sucrose metabolism of zymomonas mobilis. the levu and invb genes were clustered with a 155bp interval on the chromosome. both genes were transcribed constitutively at the basal level and the transcription of both genes was induced significantly when sucrose was added to the medium. these genes were transcribed as a bicistronic mrna and the expression was modulated by a single promoter, ...199910333527
purification and characterization of a levanbiose-producing levanase from pseudomonas sp. no. 43.a levanbiose-accumulating levanase from pseudomonas sp. no. 43 was purified to a homogeneous state by (nh4)2so4 fractionation and by chromatography on deae-toyopearl 650 m and phenyl-toyopearl 650 m columns. the molecular mass and isoelectric point of the enzyme were estimated to be 36 kda and 5.7 respectively; the optimal ph and temperature for the enzyme reaction were ph 7.0 and 40 degrees c respectively. the purified enzyme was stable in the ph range 6.0-8.0 at 20 degrees c and stable up to 5 ...199910334957
enhancement of expression and apparent secretion of erwinia chrysanthemi endoglucanase (encoded by celz) in escherichia coli b.escherichia coli b has been engineered as a biocatalyst for the conversion of lignocellulose into ethanol. previous research has demonstrated that derivatives of e. coli b can produce high levels of erwinia chrysanthemi endoglucanase (encoded by celz) as a periplasmic product and that this enzyme can function with commercial fungal cellulase to increase ethanol production. in this study, we have demonstrated two methods that improve celz expression in e. coli b. initially, with a low-copy-number ...199910347024
characterization of the tra2 region of the inchi1 plasmid this study, the dna sequence of one of the transfer regions of the inchi1 plasmid r27 was determined. this region, which corresponds to coordinates 0-40 on the r27 map has been called the tra2 region, and is believed to be involved in mating pair formation. dna sequence analysis of the transfer region identified 11 open reading frames which showed similarities to the transfer genes from other conjugative systems. the r27 transfer genes appear to most closely resemble the genes from the f plas ...199910366528
metabolic state of zymomonas mobilis in glucose-, fructose-, and xylose-fed continuous cultures as analysed by 13c- and 31p-nmr spectroscopy.the reasons for the well-known significantly different behaviour of the anaerobic, gram-negative, ethanologenic bacterium zymomonas mobilis during growth on fructose (i.e. decreased growth and ethanol yields, increased by-product formation) as compared to that on its second natural substrate, glucose, have remained unexplained. a xylose-fermenting recombinant strain of z. mobilis that was recently constructed in our laboratory also unexpectedly displayed an increased formation of by-products and ...199910369893
crystal structure of alginate lyase a1-iii from sphingomonas species a1 at 1.78 a resolution.the three-dimensional structure of alginate lyase a1-iii (alyiii) from a sphingomonas species a1 was determined by x-ray crystallography. the enzyme was crystallized by the hanging-drop vapour-diffusion method in the presence of 49% ammonium sulfate at 20 degrees c. the crystals are monoclinic and belong to the space group c2 with unit cell dimensions of a=49.18 a, b=93.08 a, c=82.10 a and beta=104.12 degrees. there was one molecule of alginate lyase in the asymmetric unit of the crystal. the di ...199910390348
development and application of a system for analysis of mixed cultures of microorganisms.development and application of a system for real-time quantitative assessment of individual cell activities in a mixed culture system was investigated. this was based on a concept that the activities of individual cells in a mixed culture can be assessed if the cells are physically separated (in separate compartments) in a vessel while the culture conditions, including the broth components, are maintained the same in all the compartments during the cultivation. on this basis, three different app ...199910394620
kinetics and performance of a co-immobilised system of amyloglucosidase and zymomonas mobilis.high operational stability and productivity of co-immobilised systems are important aspects for their successful application in industrial processes. a dynamic model is required to describe artificially co-immobilised systems because the time needed to reach steady state normally exceeds the operational life span of these systems. time dependent intraparticle concentration profiles and macroscopic conversion were modelled to study the operational stability and productivity of these systems theor ...199910397826
the efficient export of nadp-containing glucose-fructose oxidoreductase to the periplasm of zymomonas mobilis depends both on an intact twin-arginine motif in the signal peptide and on the generation of a structural export signal induced by cofactor binding.the periplasmic, nadp-containing glucose-fructose oxidoreductase of the gram-negative bacterium zymomonas mobilis belongs to a class of redox cofactor-dependent enzymes which are exported with the aid of a signal peptide containing a so-called twin-arginine motif. in this paper we show that the replacement of one or both arginine residues results in drastically reduced translocation of glucose-fructose oxidoreductase to the periplasm, showing that this motif is essential. mutant proteins which, ...199910406965
mutagenesis and crystallographic studies of zymomonas mobilis trna-guanine transglycosylase to elucidate the role of serine 103 for enzymatic activity.the trna modifying enzyme trna-guanine transglycosylase (tgt) is involved in the exchange of guanine in the first position of the anticodon with preq1 as part of the biosynthesis of the hypermodified base queuine (q). mutation of ser90 to an alanine in escherichia coli tgt leads to a dramatic reduction of enzymatic activity (reuter, k. et al. (1994) biochemistry 33, 7041-7046). to further clarify the role of this residue in the catalytic center, we have mutated the corresponding ser103 of the cr ...199910413112
a zymomonas mobilis mutant with delayed growth on high glucose concentrations.exponentially growing cells of zymomonas mobilis normally exhibit a lag period of up to 3 h when transferred from 0.11 m (2%) to 0.55 m (10%) glucose liquid medium. a mutant of z. mobilis (cu1rif2), fortuitously isolated, showed more than a 20-h lag period when grown under the same conditions, whereas on 0.55 m glucose solid medium, it failed to grow. the growth of cu1rif2 on elevated concentrations of other fermentable (0.55 m sucrose or fructose) or nonfermentable (0.11 m glucose plus 0.44 m m ...199910419959
cloning and sequencing of the beta-fructofuranosidase gene from bacillus sp. v230.the beta-fructofuranosidase gene (bff) from bacillus sp. v230 has been cloned in escherichia coli and its nucleotide sequence has been analyzed. the product of bff consists of a signal sequence of 32 amino acid (a.a.) residues for secretion and 455 a.a. residues of the extracellular beta-fructofuranosidase. the a.a. sequence of the bff product has similarities with those of the bacillus subtilis levanscrase (63.7% identity), the streptococcus mutans fructosyltransferase (33.7%), and the zymomona ...199910427700
exceptional characteristics of heterotetrameric (alpha 2 beta 2) e1p of the pyruvate dehydrogenase complex from zymomonas mobilis: expression from an own promoter and a lipoyl domain in e1 the pyruvate dehydrogenase complex (pdhc) of zymomonas mobilis the beta subunit of the pyruvate dehydrogenase (e1p) as well as the acetyltransferase (e2p) contain an n-terminal lipoyl domain. both lipoyl domains were acetylated in vitro using 2-14c-pyruvate as a substrate, demonstrating that both lipoyl domains can accept acetyl groups from the e1 component. as previously shown the structural genes (pdha alpha beta, pdhb, lpd) encoding the pyruvate dehydrogenase complex of z. mobilis are loca ...199910436929
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