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nitrite inhibition of vitreoscilla hemoglobin (vhb) in recombinant e. coli: direct evidence that vhb enhances recombinant protein production.bacteria engineered with the gene (vgb) encoding vitreoscilla hemoglobin (vhb) typically produce more protein than unengineered cells, and it has generally been assumed that vhb is responsible for this effect. here, using matched strains of e. coli that bear a recombinant alpha-amylase gene (mk57) or the alpha-amylase gene and vgb (mk79), we provide evidence supporting this assumption. sodium nitrite (which is known to inhibit heme proteins) was tested over a range of concentrations regarding ef ...200011101316
[expression of vitreoscilla hemoglobin gene in streptomyces avermitilis].two expression vectors, pwy101 and pwy102, were constructed by cloning vitreoscilla hemoglobin gene(vhb) with its native oxygen-regulated promoter into e. coli-streptomyces shuttle vector pij653. they were introduced into streptomyces avermitilis, but western blotting experiment failed to detect vhb gene expression. phz1252 is another shuttle vector for expressing vhb, in which vhb structural gene is controlled by a strong, thiostrepton-inducible streptomyces promoter ptipa. phz1252 was transfor ...200012548878
expression of alcaligenes eutrophus flavohemoprotein and engineered vitreoscilla hemoglobin-reductase fusion protein for improved hypoxic growth of escherichia coli.expression of the vhb gene encoding hemoglobin from vitreoscilla sp. (vhb) in several organisms has been shown to improve microaerobic cell growth and enhance oxygen-dependent product formation. the amino-terminal hemoglobin domain of the flavohemoprotein (fhp) of the gram-negative hydrogen-oxidizing bacterium alcaligenes eutrophus has 51% sequence homology with vhb. however, like other flavohemoglobins and unlike vhb, fhp possesses a second (carboxy-terminal) domain with nad(p)h and flavin aden ...200010618209
[expression and roles of hemoglobin gene in bacillus subtilis].hemoglobin of the gram-negative bacterium vitreoscilla can bind oxygen strongly and reduce the oxygen requirement of the bacterium. a recombinant plasmid pav was constructed by inserting the hemoglobin gene (vgb) to the downstream of the promoter of beta-lactase gene of the b. subtilis plasmid pak4. the plasmid pav was transferred into strain db104, g331 (a subtilisin-producing b. subtilis) and b53 (a xylanase-producing b. subtilis). the subclones and transfer of the vgb gene into pak4 were dete ...200010887688
error-prone pcr of vitreoscilla hemoglobin (vhb) to support the growth of microaerobic escherichia coli.expression of the gene encoding bacterial hemoglobin (vhb) from vitreoscilla has been previously used to improve recombinant cell growth and enhance product formation under microaerobic conditions. it is very likely that the properties of vhb are not optimized for foreign hosts; therefore, we used error-prone pcr to generate a number of randomly mutated vhb genes to be expressed and studied in escherichia coli. in addition, the mutated vhb proteins also contained an extension of eight residues ( ...200011005927
cloning and expression of vitreoscilla hemoglobin gene in burkholderia sp. strain dnt for enhancement of 2,4-dinitrotoluene degradation.the gene (vgb) encoding the hemoglobin (vhb) of vitreoscilla sp. was cloned into a broad host range vector and stably transformed into burkholderia (formerly pseudomonas) sp. strain dnt, which is able to degrade and metabolize 2,4-dinitrotoluene (dnt). vgb was stably maintained and expressed in functional form in this recombinant strain (yv1). when growth of yv1, in both tryptic soy broth and minimal salts broth containing dnt and yeast extract, was compared with that of the untransformed strain ...200010662485
[study on production of poly-beta-hydroxybutyrate by recombinant strain vg1(ptu14)].cloning and expression of vitreoscilla hemoglobin gene (vgb) and lambda phage lytic genes (s-rrz) in production of poly-beta-hydroxybutyrate(phb) was studied in different escherichia coli hosts such as e. colijm105, e. colijm109 and vg1. in the recombinant strains, vg1(ptu14) was a superior one which simultaneously contained three exogenes including vgb, s-rrz and phb biosynthesis genes(phbcab). the experimental results showed that after 82 hours fed-batch cult ure in lbg medium in shaking flask ...200012548993
proton nmr investigation of heme and surrounding proton in low-spin cyanide-ligated bacterial hemoglobin from vitreoscilla.(1)h nmr spectra of low-spin cyanide-ligated bacterial hemoglobin from vitreoscilla (vthb-cn) are reported. the assignments of the(1)h nmr spectra of vthb-cn have been made through mcosy, noesy, 1d toe and superweft experiments. almost all resonance peaks of heme and ligated his85 are identified. the spin-lattice relaxation time t (1)'s and the variation relationships of chemical shifts of these peaks with temperature have been acquired, from which the distances between the measured protons and ...200018763116
construction and selection of the novel recombinant escherichia coli strain for poly(beta-hydroxybutyrate) production.heterogeneous cloning of vitreoscilla hemoglobin gene (vgb), lytic genes of phage lambda with s amber mutation (s(-)rrz) and phb biosynthetic genes (phbcab) in the same host strain e. coli jm105 was carried out for production of poly(beta-hydroxybutyrate) (phb). a superior novel strain, vg1 (ptu14), was constructed and selected, which contained the vgb gene in the chromosomal dna and the plasmid ptu14 containing s(-)rrz and phbcab genes. when cultured in 100 ml of lbg medium in a 300-ml flask, a ...200016232750
construction of a novel engineering strain e.coli g830,which is adoptable to high-cell-density fermentation.a novel engineered strain g830 adoptable to high-cell-density fermentation by integrating bacterial hemoglobin vhb (vitreoscilla hemoglobin gene) into thr operon in the chromosome of pa1 blocking pta-ack metabolic pathway through the homologous recombination between the homologous fragments of integrated plasmid and that of chromosome. the engineered strain g830 was characterized by phenotype observation, pcr, thr mutant, acetate acid detection, western blotting and vhb activity assays. in high ...200112050789
optimization of immunogold labeling tem. an elisa-based method for rapid and convenient simulation of processing conditions for quantitative detection of antigen.we developed an elisa-based method for rapid optimization of various tissue processing parameters in immunogold labeling for electron microscopy. the effects of aldehyde fixation, tannic acid, postfixation, dehydration, temperature, and antigen retrieval on antibody binding activity of vitreoscilla hemoglobin (vhb) expressed in e. coli cells were assayed by elisa and the results confirmed by quantitative immunogold labeling transmission electron microscopy (tem). our results demonstrated that lo ...200111181739
[the effects of vitreoscilla hemoglobin expression on growth and antibiotic production in streptomyces cinnamonensis].streptomyces cinnamonensis a-10 is a monensin \ producing strain. phz1252 is an e. coli-streptomyces shuttle vector for expressing vhb, in which vhb structural gene is controlled by thiostrepton-inducible promoter ptipa. phz1252 was unstable in s. cinnamonensis, occurring deletion recombination. the deletion dna fragment of phz1252 included not only part of e. coli plasmid but also vhb gene. however, plasmid phz1252 isolated from s. avermitilis transformants, which lost part of e. coli plasmid b ...200111336066
characterization of an inducible oxidative stress response in vitreoscilla c1.vitreoscilla becomes resistant to killing by hydrogen peroxide and heat shock when pretreated with nonlethal levels of hydrogen peroxide. the pretreated vitreoscilla cells (60 microm hydrogen peroxide for 120 min) significantly increased survival of the lethal dose of 20 mm hydrogen peroxide or heat shock (22 degrees c --> 37 degrees c). this indicates the existence of an adaptive response to oxidative stress. however, cells pretreated with 60 microm hydrogen peroxide became nonresistant to a le ...200111355702
[cloning and expression of vhb gene in d-arabitol producing yeast].recombinant plasmid pvgb-ex2 containing vitreoscilla hemoglobin gene vgb and formaldehyde resistant gene sfa1 was constructed and transformed into d-arabitol producing yeast strain saccharomyces sp. x-62. the fact that the amount of vhb in transformant cells was considerably higher than that in control cells indicated that gene vgb was expressed in transformant cells. d-arabitol productivity and yield of fermentation by transformants were improved. the most improvement of d-arabitol productivity ...200112549085
[effect of the gene for bacterial hemoglobin vhb on the effectiveness of the process of escherichia coli-streptomyces interspecies conjugation and production of antibiotics in streptomycetes].the bacterial hemoglobin vhb gene was cloned from sliding bacterium vitreoscilla sp. as an element of the system ensuring survival of this microorganism in an environment that contains insufficient amount of oxygen. the vhb gene was transferred from escherichia coli to some streptomyces strains, producers of antibiotics, by the method of intergeneric conjugation using conjugative-integrative plasmid vectors pih1 and pch2. the stability of plasmid dna inheritance was analyzed in the genomes of ex ...200111357376
novel hemoglobins to enhance microaerobic growth and substrate utilization in escherichia coli.limited oxygen availability is a prevalent problem in microbial biotechnology. recombinant escherichia coli expressing the hemoglobin from vitreoscilla (vhb) or the flavohemoglobin from ralstonia eutropha (formerly alcaligenes eutrophus) (fhp) demonstrate significantly increased cell growth and productivity under microaerobic conditions. we identify novel bacterial hemoglobin-like proteins and examine if these novel bacterial hemoglobins can elicit positive effects similar to vhb and fhp and if ...200111587567
chromosomal integration of the vitreoscilla hemoglobin gene in burkholderia and pseudomonas for the purpose of producing stable engineered strains with enhanced bioremediating ability.using the put-minitn5 vector system developed by the laboratory of k.n. timmis, the vitreoscilla hemoglobin gene (vgb) was integrated into the chromosomes of pseudomonas aeruginosa and burkholderia cepacia; vitreoscilla hemoglobin (vhb) was expressed at 8.8 and 0.8 nmol/g wet weight of cells in the respective engineered strains. the vgb-bearing p. aeruginosa outgrew wild-type p. aeruginosa and degraded benzoic acid faster than the latter strain at both normal and low aeration. in contrast, the v ...200111598807
cell growth and oxygen uptake of escherichia coli and pseudomonas aeruginosa are differently effected by the genetically engineered vitreoscilla hemoglobin gene.vitreoscilla hemoglobin is a good oxygen trapping agent and its presence in genetically engineered escherichia coli helps this bacterium to grow better. here, the potential use of this hemoglobin, for improving the growth and the oxygen transfer properties of pseudomonas aeruginosa as well as escherichia coli, was investigated. to stably maintain it in both bacteria, a broad-host range cosmid vector (phg1), containing the entire coding sequence for vitreoscilla hemoglobin gene and its native pro ...200111164963
effects of culture conditions on enhancement of 2,4-dinitrotoluene degradation by burkholderia engineered with the vitreoscilla hemoglobin gene.growth and degradation of 2,4-dinitrotoluene (2,4-dnt) were compared in liquid cultures in shake flasks for burkholderia sp. strain dnt and strain dnt engineered to produce vitreoscilla (bacterial) hemoglobin (strain yv1). parameters varied included aeration rate, initial 2,4-dnt concentration (50 and 200 ppm), and concentration and type of cosubstrate (yeast extract, succinate, casamino acids, and tryptic soy broth). 2,4-dnt degradation increased with increasing cosubstrate concentration and wa ...200111312715
[introduction of vitreoscilla hemoglobin gene in a recombinant e. coli for phb production].vitreoscilla hemoglobin gene (vgb) was introduced into the recombinant escherichia coli vg1(ptu14) for production of poly-beta-hydroxybutyrate (phb), in order to enhance the oxygen uptake of strain from molecular level and to solve the problem of oxygen limitation in process of fermentation. the carbon monoxide difference spectra analysis of vitreoscilla hemoglobin (vhb) showed that vgb could be successfully expressed in vg1 (ptu14), and this expression was under the regulation of dissolved oxyg ...200112552801
vitreoscilla hemoglobin. intracellular localization and binding to membranes.the obligate aerobic bacterium, vitreoscilla, synthesizes elevated quantities of a homodimeric hemoglobin (vhb) under hypoxic growth conditions. expression of vhb in heterologous hosts often enhances growth and product formation. a role in facilitating oxygen transfer to the respiratory membranes is one explanation of its cellular function. immunogold labeling of vhb in both vitreoscilla and recombinant escherichia coli bearing the vhb gene clearly indicated that vhb has a cytoplasmic (not perip ...200111331274
monomer-dimer equilibrium and oxygen binding properties of ferrous vitreoscilla hemoglobin.the monomer-dimer equilibrium and the oxygen binding properties of ferrous recombinant vitreoscilla hemoglobin (vitreoscilla hb) have been investigated. sedimentation equilibrium data indicate that the ferrous deoxygenated and carbonylated derivatives display low values of equilibrium dimerization constants, 6 x 10(2) and 1 x 10(2) m(-1), respectively, at ph 7.0 and 10 degrees c. the behavior of the oxygenated species, as measured in sedimentation velocity experiments, is superimposable to that ...200111478898
dissection of central carbon metabolism of hemoglobin-expressing escherichia coli by 13c nuclear magnetic resonance flux distribution analysis in microaerobic bioprocesses.escherichia coli mg1655 cells expressing vitreoscilla hemoglobin (vhb), alcaligenes eutrophus flavohemoprotein (fhp), the n-terminal hemoglobin domain of fhp (fhpg), and a fusion protein which comprises vhb and the a. eutrophus c-terminal reductase domain (vhb-red) were grown in a microaerobic bioreactor to study the effects of low oxygen concentrations on the central carbon metabolism, using fractional (13)c-labeling of the proteinogenic amino acids and two-dimensional [(13)c, (1)h]-correlation ...200111157231
[cloning and expression of vitreoscilla hemoglobin in streptomyces aureofaciens].vitreoscilla hemoglobin gene was cloned in streptomyces aureofaciens through e. coli-streptomyces shuttle plasmids constructed by both pij699-puc19(vhb) and pij702-pbr322(vhb). under low dissolved oxygen conditions, expression of hemoglobin enhanced ctc yield of engineering strain more about 40%-60% that that of control by improving oxygen transmission in cells. hemoglobin expression by promoter of tetracycline resistance gene was more effective for oxygen transmission than that by its native pr ...200212557370
chimeric vitreoscilla hemoglobin (vhb) carrying a flavoreductase domain relieves nitrosative stress in escherichia coli: new insight into the functional role of vhb.dimeric hemoglobin (vhb) from the bacterium vitreoscilla sp. strain c1 displays 30 to 53% sequence identity with the heme-binding domain of flavohemoglobins (flavohbs) and exhibits the presence of potential sites for the interaction with its fad/nadh reductase partner. the intersubunit contact region of vhb indicates a small interface between two monomers of the homodimer, suggesting that the vhb dimers may dissociate easily. gel filtration chromatography of vhb exhibited a 25 to 30% monomeric p ...200211772621
rhizobium etli mutant modulates carbon and nitrogen metabolism in phaseolus vulgaris nodules.the aim of this study was to evaluate the biochemical events in root nodules which lead to increased yield when bean is inoculated with a rhizobium etli mutant (cfn037) having increased respiratory capacity. cfn037-inoculated plants had 22% more nitrogen (n) than did wild-type (ce3)-inoculated plants. root nodule enzymes involved in nodule carbon and nitrogen assimilation as well as in ureides and amides synthesis were assessed in plants inoculated with cfn037 and the ce3. our results show that ...200212118889
optimization of immunogold labeling tem: an elisa-based method for evaluation of blocking agents for quantitative detection of antigen.we developed an elisa-based method for rapid selection of optimal blocking agents to be used in antigen quantification by immunogold labeling electron microscopy. casein, skim milk, bsa from two sources, acetylated bsa, fish skin gelatin, horse serum, and goat serum were tested for their ability to block nonspecific binding of antibody to recombinant vitreoscilla hemoglobin (vhb) antigen expressed in escherichia coli cells by elisa and the results were confirmed by quantitative immunogold labeli ...200212019302
nitric oxide scavenging and detoxification by the mycobacterium tuberculosis haemoglobin, hbn in escherichia coli.nitric oxide (no), generated in large amounts within the macrophages, controls and restricts the growth of internalized human pathogen, mycobacterium tuberculosis h37rv. the molecular mechanism by which tubercle bacilli survive within macrophages is currently of intense interest. in this work, we have demonstrated that dimeric haemoglobin, hbn, from m. tuberculosis exhibits distinct nitric oxide dioxygenase (nod) activity and protects growth and cellular respiration of heterologous hosts, escher ...200212207698
vitreoscilla hemoglobin binds to subunit i of cytochrome bo ubiquinol oxidases.the bacterium, vitreoscilla, can induce the synthesis of a homodimeric hemoglobin under hypoxic conditions. expression of vhb in heterologous bacteria often enhances growth and increases yields of recombinant proteins and production of antibiotics, especially under oxygen-limiting conditions. there is evidence that vhb interacts with bacterial respiratory membranes and cytochrome bo proteoliposomes. we have examined whether there are binding sites for vhb on the cytochrome, using the yeast two-h ...200212080058
effect of vitreoscilla hemoglobin biosynthesis in escherichia coli on production of poly(beta-hydroxybutyrate) and fermentative parameters.in order to attain high cell density and low cost production of poly(beta-hydroxybutyrate) (phb), the vitreoscilla globin gene (vgb) was introduced into a novel recombinant strain, escherichia coli vg1 (ptu14). experiments showed that the expression of vgb was under the regulation of dissolved oxygen (do) in broth and the introduction of vgb in vg1 (ptu14) induced the parent promotion effect on cell growth and phb accumulation, especially under low do conditions. further experiments indicated th ...200212351235
pre-screening for antigen detectability in cells: a tem-based solid phase digital immunogold detection method utilizing ultra low volumes of reagents.rapid and sensitive pre-screening for the presence of antigens in cell samples and confirmation of reactivity of antibodies, before proceeding with electron microscopy, is highly desirable. most of the methods developed for this purpose are generally not very efficient and suitable for dealing with very small volumes of sample and reagents. in this work we present a simple, sensitive and rapid solid phase transmission electron microscope (tem) based method for the detection of picogram (pg) leve ...200212423260
[hemoglobin, from microorganisms to man: a single structural motif, multiple functions].haemoglobins from unicellular organisms, plants or animals, share a common structure, which results from the folding, around the heme group, of a polypeptide chain made from 6-8 helices. nowadays, deciphering the genome of several species allows one to draw the evolutionary tree of this protein going back to 1800 millions of years, at a time when oxygen began to accumulate in the atmosphere. this permits to follow the evolution of the ancestral gene and of its product. it is likely that, only in ...200212520866
[study on effect of vitreoscilla hemoglobin gene expression on growth and metabolism of streptomyces aureofaciens].vitreoscilla hemoglobin gene was expressed in s. aureofaciens through promoter of tetracycline resistance gene. characteristics of s. aureofaciens growth and metabolism were studied in 1 m3 fermenter. in high dissolved oxygen concentrations, expression of hemoglobin gene had little effect on growth and metabolism of s. aureofaciens, and there were no obvious differences between engineering strain and control. the chlortetracycline of engineering strain was 22905 u/ml and of control was 22896 u/m ...200212557546
purification, crystallization and preliminary x-ray analysis of the soluble domain of the na+-pumping cytochrome bo quinol oxidase from vitreoscilla.the 24 kda cyoa soluble domain of vitreoscilla cytochrome bo quinol oxidase, which pumps out na(+) during respiration, has been crystallized from a solution of 2 m ammonium sulfate and 5% 2-propanol. the crystal belongs to cubic space group p4(3)32, with unit-cell parameters a = b = c = 122.2 a, alpha = beta = gamma = 90 degrees and one subunit in the asymmetric unit. a 99.8% complete data set to 3.3 a has been collected at the 17-id beamline of the advanced photon source. the structure was dete ...200212136145
improved cell growth in tobacco suspension cultures expressing vitreoscilla hemoglobin.expression of the gene encoding bacterial hemoglobin (vhb) from vitreoscilla has been previously used to improve recombinant cell growth and enhance product formation under microaerobic conditions, a common phenomenon in large-scale cultivations of bacteria. this technology has now been applied to tobacco suspension cultures. tobacco suspension cultures have been generated from vhb-expressing tobacco plants. cell cultures were capable of producing an active hemoglobin. when grown in shake flasks ...200211934289
the x-ray structure of ferric escherichia coli flavohemoglobin reveals an unexpected geometry of the distal heme pocket.the x-ray structure of ferric unliganded lipid-free escherichia coli flavohemoglobin has been solved to a resolution of 2.2 a and refined to an r-factor of 19%. the overall fold is similar to that of ferrous lipid-bound alcaligenes eutrophus flavohemoglobin with the notable exception of the e helix positioning within the globin domain and a rotation of the nad binding module with respect to the fad-binding domain accompanied by a substantial rearrangement of the c-terminal region. an inspection ...200211964402
intracellular expression of vitreoscilla hemoglobin modifies microaerobic escherichia coli metabolism through elevated concentration and specific activity of cytochrome o.the function of the reversible oxygen-binding hemoprotein from vitreoscilla (vhb), which enhances oxygen-limited cell growth and recombinant protein production when functionally expressed in escherichia coli, was investigated in wild-type e. coli and in e. coli mutants lacking one of the two terminal oxidases, cytochrome o complex (aerobic terminal oxidase, cyo) or cytochrome d complex (microaerobic terminal oxidase, cyd). deconvolution of vhb, cytochrome o, and cytochrome d bands from in vivo a ...200212209827
comparison of green fluorescent protein expression in two industrial escherichia coli strains, bl21 and w3110, under co-expression of bacterial hemoglobin.vitreoscilla hemoglobin (vhb) has been successfully used to enhance production of foreign proteins in several microorganisms including escherichia coli. we compared the expression of an oxygen-dependent foreign protein, green fluorescent protein (gfp) under co-expression of vhb in two typical industrial e. coli strains, bl21 (a b derivative) and w3110 (a k12 derivative), which have different metabolic properties. we employed the nar oxygen-dependent promoter for self-tuning regulation of vhb exp ...200212172620
inverse metabolic engineering: a strategy for directed genetic engineering of useful phenotypes.the classical method of metabolic engineering, identifying a rate-determining step in a pathway and alleviating the bottleneck by enzyme overexpression, has motivated much research but has enjoyed only limited practical success. intervention of other limiting steps, of counter-balancing regulation, and of unknown coupled pathways often confounds this direct approach. here the concept of inverse metabolic engineering is codified and its application is illustrated with several examples. inverse me ...200212209828
bacterial hemoglobins and flavohemoglobins for alleviation of nitrosative stress in escherichia coli.escherichia coli mg1655 cells expressing novel bacterial hemoglobin and flavohemoglobin genes from a medium-copy-number plasmid were grown in shake flask cultures under nitrosative and oxidative stress. e. coli cells expressing these proteins display enhanced resistance against the no(.) releaser sodium nitroprusside (snp) relative to that of the control strain bearing the parental plasmid. expression of bacterial hemoglobins originating from campylobacter jejuni (chb) and vitreoscilla sp. (vhb) ...200212324328
expression of double vitreoscilla hemoglobin enhances growth and alters ribosome and trna levels in escherichia coli.in several organisms, expression of a gene encoding dimeric hemoglobin (vhb) from the obligate aerobic bacterium vitreoscilla stercoraria has been shown to increase microaerobic cell growth and enhance oxygen-dependent cell metabolism. in an attempt to further improve these effects of vhb, a gene encoding two vhb genes connected by a short linker of six base pairs was constructed and expressed in escherichia coli(double vhb). escherichia coli cells expressing double vhb reached a cell density 19 ...200212052087
isolation, sequencing, and characterization of the cytochrome bo operon from vitreoscilla.the entire operon encoding the sodium pumping cytochrome bo from the bacterium vitreoscilla was isolated and sequenced, and this sequence was analyzed by blast and hydropathy plots. there are fairly similar phylogenetic relationships which apply to all five proteins, but overall greater similarity to members of the gamma subdivision than the beta subdivision of the proteobacteria. hydropathy plots of all five cyo proteins show near identity with those of the corresponding e. coli subunits, indic ...200312751331
effects of vitreoscilla hemoglobin on the 2,4-dinitrotoluene (2,4-dnt) dioxygenase activity of burkholderia and on 2,4-dnt degradation in two-phase bioreactors.expression of vgb, encoding vitreoscilla hemoglobin (vhb), in burkholderia strain yv1 was previously shown to improve cell growth and enhance 2,4-dinitrotoluene (2,4-dnt) degradation compared with control strain dnt, especially under hypoxic conditions. in the work reported here, the ratio of 2,4-dnt degraded to oxygen uptake was approximately 5-fold larger for strain yv1 than for strain dnt. the addition of purified vhb to cytosolic fractions of strain dnt increased 2,4-dnt degradation 1.5-fold ...200312743828
menadione knocks out vitreoscilla haemoglobin (vhb): the current evidence for the role of vhb in recombinant escherichia coli.genetically engineering the heterologous bacterial host with the gene (vgb) encoding vitreoscilla haemoglobin (vhb) has been found to provide typical advantages in growth and production, and it has generally been assumed that vhb is responsible for this effect. here, using matched strains of escherichia coli that bear a recombinant r-amylase gene (mk57) or the r-amylase gene and vgb (mk79), we examined this assumption. menadione, which is known to oxidize haem proteins, was tested over a range o ...200312659636
improving heterologous gene expression in aspergillus niger by introducing multiple copies of protein-binding sequence containing ccaat to the promoter.to increase the expression level of heterologous gene in aspergillus niger.200312753242
flavohemoglobin hmp, but not its individual domains, confers protection from respiratory inhibition by nitric oxide in escherichia coli.escherichia coli possesses a two-domain flavohemoglobin, hmp, implicated in nitric oxide (no) detoxification. to determine the contribution of each domain of hmp toward no detoxification, we genetically engineered the hmp protein and separately expressed the heme (hd) and the flavin (fd) domains in a defined hmp mutant. expression of each domain was confirmed by western blot analysis. co-difference spectra showed that the hd of hmp can bind co, but the co adduct showed a slightly blue-shifted pe ...200312826671
direct electrochemical characterization of vitreoscilla sp. hemoglobin entrapped in organic films.the redox properties of a prokaryotic, vitreoscilla sp. hemoglobin (vhb) in fuzzy organic films are studied with electrochemistry. this vhb exhibits irreversible electrochemical response at bare pyrolytic graphite (pg) electrode surfaces. however, upon being entrapped in organic films, the heterogeneous electron transfer rate of vhb will be sufficiently high to produce a quasi-reversible electrochemical response. the observation of electrocatalysis (reduction of o2) by hemes suggests that the pr ...200312878030
vitreoscilla hemoglobin promoter is not responsive to nitrosative and oxidative stress in escherichia coli.the vitreoscilla hemoglobin gene (vhb) is expressed under oxygen-limited conditions via an fnr-dependent mechanism. furthermore, camp-crp has been implicated in its regulation. recently, vhb protein has been reported to protect a heterologous host from nitrosative stress. in this study we analyzed the regulation of the vitreoscilla hemoglobin promoter (pvhb) in escherichia coli under nitrosative and oxidative stress conditions. our results show unambiguously that expression of neither vhb nor ch ...200312855179
a solid-phase method for evaluation of gold conjugate used in quantitative detection of antigen by immunogold-labeling electron microscopy.rapid and sensitive screening for confirming the reactivity of reagents, before proceeding for electron microscopy, is highly desirable. elisa-based methods have been shown to be highly efficient and successful for rapid prescreening and optimization of immunological as well as sample-processing reagents for the sensitive detection and quantitation of antigen by electron microscopy. the drawback of these methods lies in their inability to provide any information regarding the gold conjugate used ...200312969545
bacterial hemoglobins and flavohemoglobins: versatile proteins and their impact on microbiology and biotechnology.in response to oxygen limitation or oxidative and nitrosative stress, bacteria express three kinds of hemoglobin proteins: truncated hemoglobins (tr hbs), hemoglobins (hbs) and flavohemoglobins (flavo hbs). the two latter groups share a high sequence homology and structural similarity in their globin domain. flavohemoglobin proteins contain an additional reductase domain at their c-terminus and their expression is induced in the presence of reactive nitrogen and oxygen species. flavohemoglobins ...200314550944
microbial globins.globins are an ancient and diverse superfamily of proteins. the globins of microorganisms were relatively ignored for many decades after their discovery by warburg in the 1930s and rediscovery by keilin in the 1950s. the relatively recent focus on them has been fuelled by recognition of their structural diversity and fine-tuning to fulfill (probably) discrete functions but particularly by the finding that a major role of certain globins is in protection from the stresses caused by exposure to ni ...200314560666
expression of vitreoscilla hemoglobin improves growth and levels of extracellular enzyme in yarrowia lipolytica.enhancement in oxygen uptake by high-cell-density cultivations has been achieved previously by expression of the bacterial hemoglobin gene from vitreoscilla. the vitreoscilla hemoglobin (vhb) gene was expressed in the yeast yarrowia lipolytica to study the effect of expression in this commercially important yeast. the expression of vhb in this yeast was found to enhance growth, contrary to reported observations in wild-type saccharomyces cerevisiae in which there was no significant growth enhanc ...200314595778
biodegradation of 2-chlorobenzoate by recombinant burkholderia cepacia expressing vitreoscilla hemoglobin under variable levels of oxygen availability.the influence of bacterial hemoglobin, vhb, on dechlorination and degradation of 2-chlorobenzoate (2-cba) by recombinant burkholderia sp. under variable oxygen availability with an initial dissolved oxygen concentration of 0.27 mm-0.72 mm was investigated in batch and continuous culture. ability to express vhb was provided to recombinant burkholderia by transformation with the vhb gene, vgb, on plasmid psc160. 100% of 0.5 mm cba was degraded in cultures with 85% and 70% of total volume as headsp ...200314571952
fusion protein system designed to provide color to aid in the expression and purification of proteins in escherichia coli.we have designed and constructed a new fusion expression vector (pkw32), which contains the his-tagged vitreoscilla hemoglobin (vhb) coding gene upstream of the multiple cloning site. the pkw32 vector was designed to express target proteins as vhb fusions, which can be purified in one step by affinity chromatography. due to the color of the heme in vhb, the vhb-fused target proteins have a red color that provides a visual aid for estimating their expression level and solubility. the red color ca ...200314597006
fusion protein of vitreoscilla hemoglobin with d-amino acid oxidase enhances activity and stability of biocatalyst in the bioconversion process of cephalosporin c.in this study we constructed an artificial flavohemoprotein by fusing vitreoscilla hemoglobin (vhb) with d-amino acid oxidase (dao) of rhodotorula gracilis to determine whether bacterial hemoglobin can be used as an oxygen donor to immobilized flavoenzyme. this chimeric enzyme significantly enhanced dao activity and stability in the bioconversion process of cephalosporin c. in a 200-ml bioreactor, the catalytic efficiency of immobilized vhb-dao against cephalosporin c was 12.5-fold higher than t ...200312632405
correlation between bacterial haemoglobin gene (vgb) and aeration: their effect on the growth and alpha-amylase activity in transformed enterobacter aerogenes.to evaluate the effects of bacterial haemoglobin on bacterial growth and alpha-amylase formation under different aeration conditions.200312752814
enhanced ribosome and trna contents in escherichia coli expressing a truncated vitreoscilla hemoglobin mutant analyzed by flow field-flow fractionation.the ribosome and trna levels of escherichia coli cells, transformed with a native or mutated vitreoscilla hemoglobin genes (vhb), were investigated using asymmetrical flow field-flow fractionation (affff). mutagenesis of rhb by error-prone pcr was carried out to alter the growth behavior of microaerobically cultivated native vhb-expressing e. coli. a vhb mutant, pvmt1, was identified, which was able to reach a remarkably high final a600 of 15, the value of which being 160% higher than that of a ...200314571972
expression of vitreoscilla haemoglobin in hybrid aspen (populus tremula x tremuloides).we describe the first ever expression of vitreoscilla haemoglobin (vhb) in an economically important boreal woody plant hybrid aspen (populus tremula x tremuloides). vhb has mainly been expressed in biotechnologically important unicellular organisms of both prokaryotic and eukaryotic origin. vhb expression, in this study, was analysed under different greenhouse cultivation conditions and under elevated uv-b illumination. microscope analyses of leaves grown under optimized conditions revealed sig ...200317163905
[recent advances in the study of vitreoscilla hemoglobin(vhb) and related proteins - cloning, expression and physiological actions in heterologons hosts and in transgenic tobacco plants].the first bacterial hemoglobin(vhb) was found in a strictly aerobic bacterium, vitreoscilla strain c1, occurring in marshes low in oxygen, but rich in organic matter. the hemoglobin gene is induced under low oxygen tension and may amount to 20 times as high. the expression of vhb promotes cell growth, protein biosynthesis and primary and secondary metabolism of the host cells, because the increased intracellular oxygen accelerates both the function of respiratory chain and terminal oxidases. the ...200315969050
expression of the gene coding for bacterial hemoglobin improves beta-galactosidase production in a recombinant pichia pastoris.genes coding for vitreoscilla hemoglobin (vhb) with peroxisome targeting signal (pts1) tag and beta-galactosidase were co-expressed in pichia pastoris under the alcohol oxidase1 (aox1) promoter. the expression of vhb-pts1 had no positive effect on cell growth but significantly enhanced the whole cell beta-galactosidase activity to 4-fold higher than that of vhb-free cell in yeast extract/peptone/methanol medium under aerobic cultivation.200314514050
extreme uv-a sensitivity of the filamentous gliding bacterium vitreoscilla stercoraria.strains of the filamentous gliding bacterium vitreoscilla, lb13 and c1, are shown to be highly sensitive to uv-a (320-400 nm), with an ld50 of less than 20 kj m(-2). vitreoscilla lb13 can be protected from uv-a by including superoxide dismutase and catalase, separately or in combination, during the exposure, indicating an involvement of reactive oxygen species. lb13a, a photo-insensitive strain derived from lb13, is described.200312855170
genetic engineering of enterobacter aerogenes with the vitreoscilla hemoglobin gene: cell growth, survival, and antioxidant enzyme status under oxidative stress.hemoglobins in unicellular organisms, like the one here in the bacterium vitreoscilla, have greater chemical reactivity than their homologues in multicellular organisms. they can catalyze redox reactions and may protect cells against oxidative stress. the ability of vitreoscilla hemoglobin to complement deficiencies of terminal cytochrome oxidases in escherichia coli also suggests that this hemoglobin can receive electrons during respiration. in this study, a recombinant strain of enterobacter a ...200312892849
evidence for stable transformation of pseudomonas aeruginosa with a puc-based plasmid.pseudomonas aeruginosa was transformed with puc8:16, a puc-based plasmid bearing the gene (vgb) encoding vitreoscilla (bacterial) hemoglobin (vhb). transformation was initially indicated by an increase in ampicillin resistance from 1500 to 2500 mg l(-1). presence of the plasmid in p. aeruginosa was confirmed by amplification of a portion of vgb from and detection of vhb in the transformant but not the untransformed host. southern blot analysis further indicated that puc8:16 existed as an autonom ...200312889831
vitreoscilla hemoglobin overexpression increases submergence tolerance in cabbage.agrobacterium tumefaciens was used to deliver the vhb gene into cabbage (brassica oleracea var. cabitata) cv. xiaguang's parent line, 103. using hypocotyls and cotyledon petioles as explants for infection, we obtained a transformation efficiency of 3-5% based on the number of transgenic shoots produced from the number of explants used for infection. molecular analysis indicated that the vhb gene was stably integrated into the cabbage genome and that the vhb gene was expressed at the rna level. c ...200415747159
enhanced kinetics of genetically engineered burkholderia cepacia: the role of vgb in the hypoxic metabolism of 2-cba.application of vitreoscilla hemoglobin (vhb) technology to 2-cba degradation by burkholderia cepacia strain dnt under hypoxic conditions was studied in continuous culture chemostats. dechlorination abilities of both recombinant (vhb gene (vgb) containing) and untransformed cells were investigated at various dilution rates to ensure complete degradation of 2-cba. as the dilution rate increased from 0.025 to 0.25 h(-1), the ratios of chloride release to degraded 2-cba concentration decreased from ...200415211495
enhancement of 2,4-dinitrotoluene biodegradation by burkholderia sp. in sand bioreactors using bacterial hemoglobin technology.continuous flow sand column bioreactor experiments were conducted to investigate the effect of 2,4-dinitrotoluene (dnt) concentration (i.e. dnt loading rate) and influent dissolved oxygen (do) concentration on aerobic biodegradation of dnt by wild type (dnt) and recombinant (yv1) burkholderia sp., the latter containing plasmid psc160 which carries the gene (vgb) encoding the hemoglobin (vhb) from the bacterium vitreoscilla. the experiments were conducted in two continuous flow packed bed sand co ...200415228074
cloning and expression of the vitreoscilla hemoglobin gene in enterobacter aerogenes: effect on cell growth and oxygen uptake.the hemoglobins found in unicellular organisms show a greater chemical reactivity, protect cells against oxidative stress and hence have been implicated in a wider variety of potential functions than those traditionally associated with animal and plant hemoglobins. there are well-documented studies showing that bacteria expressing vitreoscilla hemoglobin (vhb), the first prokaryotic hemoglobin characterized, have better growth and oxygen uptake rates than vhb counterparts.200415283330
enhanced production of acetoin and butanediol in recombinant enterobacter aerogenes carrying vitreoscilla hemoglobin gene.microbial production of butanediol and acetoin has received increasing interest because of their diverse potential practical uses. although both products are fermentative in nature, their optimal production requires a low level of oxygen. in this study, the use of a recombinant oxygen uptake system on production of these metabolites was investigated. enterobacter aerogenes was transformed with a puc8-based plasmid carrying the gene (vgb) encoding vitreoscilla (bacterial)hemoglobin (vhb). the pre ...200415309606
production of l-asparaginase in enterobacter aerogenes expressing vitreoscilla hemoglobin for efficient oxygen uptake.this study is the first utilizing vitreoscilla hemoglobin in a heterologous bacterium, enterobacter aerogenes, to determine the effect of such a highly efficient oxygen-uptake system on the production of l-asparaginase, an enzyme that has attracted considerable attention due to its anti-tumor activity. here, we show that the vitreoscilla hemoglobin expressing strain has from 10-fold to more than two orders of magnitude lower l-asparaginase activity than the wild type or the control without the v ...200414593509
coexpression of vitreoscilla hemoglobin reduces the toxic effect of expression of d-amino acid oxidase in e. coli.expression of the gene (daao) encoding d-amino acid oxidase (daao) in escherichia coli typically results in a marked decrease of cell viability, and it has generally been assumed that the consumption of intracellular d-alanine by daao is responsible for this effect. vitreoscilla hemoglobin (vhb) gene (vgb) was coexpressed with rhodosporidium toruloides d-amino acid oxidase in e. coli bl21(de3) and bl21(de3)plyss, expression hosts differing in the stringency of suppressing basal transcription. no ...200415458318
gene expression profiling of escherichia coli expressing double vitreoscilla haemoglobin.in a recent investigation, expression of a double vitreoscilla haemoglobin (two fused vhb molecules) in escherichia coli grown in shake flasks resulted in higher final cell density and considerably higher levels of ribosomes and trna. in this study, we have investigated the e. coli transcriptome in cells expressing native vhb, double vhb and control cells lacking vhb by hybridising mrna from the different constructs to high-density oligonucleotide arrays. within the 95% confidence interval, 4 an ...200415464604
metabolic engineering for the production of copolyesters consisting of 3-hydroxybutyrate and 3-hydroxyhexanoate by aeromonas hydrophila.aeromonas hydrophila 4ak4 was able to synthesize copolyesters consisting of 3-hydroxybutyrate (3hb) and about 15 mol-% 3-hydroxyhexanoate (3hhx) (phbhhx) when grown in long chain fatty acids such as dodecanoate regardless of growth conditions. to regulate the unit fraction in phbhhx, phba and phbb genes encoding beta-ketothiolase and acetoacetyl-coa reductase in ralstonia eutropha, were introduced into a. hydrophila 4ak4. when gluconate was used as cosubstrate of dodecanoate, the recombinant pro ...200415468215
structure-function studies of the vitreoscilla hemoglobin d-region.the d-region connecting helices c and e of vitreoscilla hemoglobin (vhb) appears disordered in the crystal structure. six site-directed mutants in this region were made to investigate its possible functions. the mutant vhb's were analyzed using uv-visible and ftir spectroscopy, using primarily the co liganded forms, and their heme/protein ratios were determined. the results implicate asp44, arg47, and glu49 as especially important in heme-globin interactions and ligand binding, and enabled const ...200415044098
molecular cloning of polyhydroxyalkanoate synthesis operon from aeromonas hydrophila and its expression in escherichia coli.the polyhydroxyalkanoate synthesis operon was cloned from aeromonas hydrophila cgmcc 0911. heterogeneous expression of the cloned polyhydroxyalkanoate synthesis operon in escherichia coli resulted in accumulation of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) consisting of 13.9 mol % 3-hydroxyhexanoate up to 29.2 wt % of cell dry weight when grown in lauric acid. the cell dry weight of recombinant e. coli harboring the polyhydroxyalkanoate synthesis operon was improved to 1.7 g l (-1), which w ...200415458314
[expression of vitreoscilla hemoglobin and nitrilase in the yeast pichia pastoris].the expression of the vgb gene in vivo could improve the fermentation density and then contribute the extracellular secretion of the product of bxn gene. constructed the recombination plasmid ppic9k-vgbbxn and transformed into pichia pastoris gs115. the results of pcr and sds-page indicate that the vgb gene and bxn gene had integrated into the genome of pichia pastoris gs115 and expressed in efficient level. also, the protein activity of their products had been verified respectively. shake flask ...200415973999
[enhanced ergosterol production by recombinant saccharomyces cerevisiae 1190 harboring vitreoscilla hemoglobin gene (vgb)].ergosterol is a principal sterol of fungi. it is a raw material for production of vitamin d2, hydrocortisone, progesterone and brassinolide. synthesis of ergosterol requires molecular oxygen, and low oxygen tensions was reported to dramatically reduce ergosterol concentration. vitreoscilla hemoglobin gene (vgb), a homodimeric hemoglobin gene from gram-negative obligate aerobic bacterium vitreoscilla, enables a higher specific cellular oxygen uptake rate, it also improves the oxygen transportatio ...200415971621
co-expression of bacterial hemoglobin overrides high glucose-induced repression of foreign protein expression in escherichia coli w3110.co-expression of vitreoscilla hemoglobin (vhb) can enhance production of foreign proteins in several microorganisms, including escherichia coli. production of foreign proteins [green fluorescent protein (gfp) and organophosphorous hydrolase (oph)] has been examined in two typical industrial e. coli strains, w3110 (a k12 derivative) and bl21 (a b derivative). in particular, we investigated the effects of vhb co-expression and media glucose concentration on target protein production. we employed t ...200415266126
expression of vitreoscilla haemoglobin in tobacco cell cultures relieves nitrosative stress in vivo and protects from no in vitro.targeted expression of vitreoscilla haemoglobin (vhb) has been analysed in nicotiana tabacum plants and suspension cultures under various growth and stress conditions. vhb localization to different cell compartments (cytoplasm, chloroplast and mitochondria) was successful, as judged by signal peptide cleavage. the presence of vhb in subcellular compartments did not result in phenotypical differences between these plant lines. in contrast with previous reports, we were unable to discern any signi ...200417147613
intracellular expression of vitreoscilla hemoglobin in aspergillus terreus to alleviate the effect of a short break in aeration during culture.a continuous supply of o(2) is important for itaconic acid production in aspergillus terreus. any interruption of aeration significantly reduces itaconic acid production. to overcome this effect, a. terreus m8 was transformed with the vitreoscilla hemoglobin gene (vgb) which, as shown by southern hybridization, was integrated into the recipient chromosome. the activity of the expressed hemoglobin was confirmed by a co-difference spectrum. during itaconic acid production, the effect of a break in ...200415218381
vitreoscilla hemoglobin renders enterobacter aerogenes highly susceptible to heavy metals.when expressed in heterologous microorganisms vitreoscilla hemoglobin (vhb) acts as oxygen storage and causes a higher oxygen uptake. in this study, the effect of this protein on growth, sensitivity and antioxidant properties of enterobacter aerogenes exposed to metal stress was investigated. the strain expressing vhb was more sensitive to mercury and cadmium as the minimal inhibitory concentration (mic) for these metals was up to 2-fold lower in this strain than the host and the recombinant str ...200415689114
role of an inducible single-domain hemoglobin in mediating resistance to nitric oxide and nitrosative stress in campylobacter jejuni and campylobacter coli.campylobacter jejuni expresses two hemoglobins, each of which exhibits a heme pocket and structural signatures in common with vertebrate and plant globins. one of these, designated cgb, is homologous to vgb from vitreoscilla stercoraria and does not possess the reductase domain seen in the flavohemoglobins. a cgb-deficient mutant of c. jejuni was hypersensitive to nitrosating agents (s-nitrosoglutathione [gsno] or sodium nitroprusside) and a nitric oxide-releasing compound (spermine nonoate). th ...200415292134
nitric oxide detoxification--a new era for bacterial globins in biotechnology?for more than a decade, the expression of vitreoscilla hemoglobin (vhb) has been used to improve the growth and/or productivity of various organisms that are important for the production of valuable metabolites and recombinant proteins by biotechnological processes. extensive experimental data have shown that vhb enhances the energy status of the cell under oxygen-limited conditions, presumably by improving the supply of intracellular oxygen. recently, bacterial globin proteins have gained more ...200515661342
antioxidant status, alpha-amylase production, growth, and survival of hemoglobin bearing escherichia coli exposed to hypochlorous acid.in the present work, two matched strains of e. coli that bear a recombinant r-amylase gene (mk57) or the r-amylase gene and vgb (mk79-hemoglobin expressing strain) were exposed to hocl. in these cells, glutathione (gsh), superoxide dismutase (sod), catalase (cat), alpha-amylase production, growth and lethality were assessed in the presence and absence of hocl. it was observed that the hemoglobin makes cells highly susceptible to killing by hocl. the maximum survival for both strains was with sta ...200516417460
expression of bacterial hemoglobin in the yeast, pichia pastoris, with a low o2-induced promoter.the psadh2-promoter of pichia stipitis alcohol dehydrogenase ii (adh ii) gene was employed to control the expression of vitreoscilla hemoglobin (vhb) gene in pichia pastoris. as in p. stipitis, the promoter was also induced microaerobically in p. pastoris. the expression level of vhb in p. pastoris at low o2 tension (<5% air saturation) was 16 nmol/g dry cell wt, i.e. about 24-fold higher than that at 60% air saturation. the expressed vhb enhanced growth of p. pastoris under microaerobic conditi ...200516231222
[current research status of vitreoscilla hemoglobin and the prospective application in traditional chinese medicine]. 200516159562
mutational study of the bacterial hemoglobin distal heme pocket.ligand binding experiments on three mutants in the distal heme pocket of vitreoscilla hemoglobin (glne7his, proe8ala, and glne7his,proe8ala) were used to probe the role of glne7 and proe8 in the pocket's unusual structure. the oxygen dissociation constants for the wild type, e8ala mutant, and e7his mutant proteins were 4.5, 4.7, and 1.7microm, respectively; the k(d) for the double mutant was not determinable by our technique. visible-soret spectra of the carbonyl and cyanyl forms and ft-ir of th ...200515582576
expression of vitreoscilla hemoglobin enhances growth of hyoscyamus muticus hairy root cultures.the vitreoscilla hemoglobin gene (vhb) was introduced into hyoscyamus muticus with the aim of investigating its effect on growth and alkaloid production of agrobacterium rhizogenes-induced hairy root cultures. we were able to generate several vhb-expressing hairy root lines with different integration patterns. substantial somaclonal variation was observed in growth and hyoscyamine production amongst both vhb-expressing lines and controls. despite this variation, the growth properties of single l ...200515678373
arca works with fnr as a positive regulator of vitreoscilla (bacterial) hemoglobin gene expression in escherichia coli.low oxygen induction of the bacterial (vitreoscilla) hemoglobin gene (vgb) by the arc system was investigated, as the presumptive vgb crp site was found to have 73% identity to the escherichia coli consensus arca site. the role of arca by itself and with fnr was examined in e. coli using the wild type vgb promoter and promoter mutants affecting the fnr and crp (presumptive arca) sites and e. coli strains with all combinations of fnr+/fnr-, arca+/arca- genotypes. high-level transcription required ...200516255146
construction of highly efficient e. coli expression systems containing low oxygen induced promoter and partition region.a series of high-copy-number escherichia coli expression vectors equipped with an oxygen-sensitive promoter p(vgb) of vitreoscilla hemoglobin (encoded by the vgb gene) were constructed and characterized. plasmid pkvp containing p(vgb) was inducible by low oxygen tension, while plasmid pkvpp containing a partition (par) region from plasmid psc101 ligated to p(vgb) provided inheritable stability for the vectors in the absence of ampicillin. plasmid pkvpv had the vitreoscilla hemoglobin operon vgb ...200515711794
improvement of bioremediation by pseudomonas and burkholderia by mutants of the vitreoscilla hemoglobin gene (vgb) integrated into their chromosomes.using genetic engineering, the vitreoscilla (bacterial) hemoglobin gene (vgb) was integrated stably into the chromosomes of pseudomonas aeruginosa and burkholderia sp. strain dnt. this was done for both wild type vgb and two site-directed mutants of vgb that produce vitreoscilla hemoglobin (vhb) with lowered oxygen affinities; in all cases functional vhb was expressed. similar to previous results, the wild type vhb improved growth for both species and degradation of 2,4-dinitrotoluene (burkholde ...200515806390
evidence that na+-pumping occurs through the d-channel in vitreoscilla cytochrome bo.the operon (cyo) encoding the na(+)-pumping respiratory terminal oxidase (cytochrome bo) of the bacterium vitreoscilla was transformed into escherichia coli gv100, a deletion mutant of cytochrome bo. this was done for the wild type operon and five mutants in three conserved cyo subunit i amino acids known to be crucial for h(+) transport in the e. coli enzyme, one near the nuclear center, one in the k-channel, and one in the d-channel. co-binding, nadh and ubiquinol oxidase, and na(+)-pumping ac ...200515910742
chimeric antibody-binding vitreoscilla hemoglobin (vhb) mediates redox-catalysis reaction: new insight into the functional role of vhb.experimentation was initiated to explore insight into the redox-catalysis reaction derived from the heme prosthetic group of chimeric vitreoscilla hemoglobin (vhb). two chimeric genes encoding chimeric vhbs harboring one and two consecutive sequences of fc-binding motif (z-domain) were successfully constructed and expressed in e. coli strain tg1. the chimeric zvhb and zzvhb were purified to a high purity of more than 95% using igg-sepharose affinity chromatography. from surface plasmon resonance ...200616967102
metabolic engineering of aeromonas hydrophila for the enhanced production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate).wild-type aeromonas hydrophila 4ak4 produced 35-45 wt.% poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (phbhhx) consisting of 10-15 mol% 3-hydroxyhexanoate (3hhx). to enhance phbhhx production, vgb gene encoding vitreoscilla haemoglobin or fadd gene encoding escherichia coli acyl-coa synthase was co-expressed with polyhydroxyalkanoates (pha) synthesis-related genes including phbab from wautersia eutropha and phapcj from a. hydrophila. expression of vgb increased phbhhx content from 46 to 53 wt.% ...200615983806
chromosomal integration of the vitreoscilla hemoglobin gene and its physiological actions in tremella fuciformis.the vitreoscilla hemoglobin (vhb) gene was expressed in yeast-like conidia (ylcs) of tremella fuciformis (t. fuciformis) to increase cell density in submerged fermentation by enhancing oxygen uptake. with the intention of doing this, an integrated expression vector containing the vhb gene and the hygromycin b phosphotransferase (hph) gene derived from escherichia coli (e. coli) as the selectable marker was constructed, and then transformed into protoplasts of ylcs from t. fuciformis with restric ...200616501972
interaction of vitreoscilla hemoglobin with membrane lipids.the interaction of the recombinant hemoglobin from vitreoscilla sp. (vhb) with the bacterial membrane of escherichia coli cells has been investigated by measuring the propensity of vhb to interact with monolayers formed by natural bacterial phosholipids. the measurements showed that the protein is capable of penetrating the monolayers, possibly establishing interactions with the hydrophobic acyl chains. vhb is also capable of binding reversibly phospholipids and free fatty acids in solution with ...200616566580
enhancement of cellulose pellicle production by constitutively expressing vitreoscilla hemoglobin in acetobacter xylinum.vitreoscilla hemoglobin (vhb) gene driven by the constitutive bla promoter was expressed in the cellulose-producing acetobacter xylinum. the expressed vhb was biochemically active and could enhance cell growth in a shaken culture containing cellulase. vhb-expressing a. xylinum (vhb+) exhibited a specific growth rate 50% higher than that of the host strain (vhb-). probably because of its faster growth rate, the size of tentacled cellulose beads produced by vhb+ was about 20% of that produced by v ...200617137307
correlation between bacterial hemoglobin and carbon sources: their effect on copper uptake by transformed e. coli strain alpha dh5.escherichia coli alphadh5 harboring vitreoscilla hemoglobin gene, vgb (escherichia:puc8:16), and control strains (parental and vgb-lacking transformed strains, e. coli:puc9) were employed for studying the potential ability of copper uptake. the strains were grown on a minimal medium, 5xm9, that contained different carbon (mannitol, fructose, lactose, and maltose) and nitrogen (casein and yeast extract) sources. a clear type-response relationship between the level of copper uptake and the type of ...200616392001
expression of vitreoscilla hemoglobin in gordonia amarae enhances biosurfactant production.the gene (vgb) encoding vitreoscilla (bacterial) hemoglobin (vhb) was electroporated into gordonia amarae, where it was stably maintained, and expressed at about 4 nmol vhb g(-1) of cells. the maximum cell mass (od(600)) of vgb-bearing g. amarae was greater than that of untransformed g. amarae for a variety of media and aeration conditions (2.8-fold under normal aeration and 3.4-fold under limited aeration in rich medium, and 3.5-fold under normal aeration and 3.2-fold under limited aeration in ...200616491354
heterologous expression of vitreoscilla hemoglobin (vhb) and cultivation conditions affect the alkaloid profile of hyoscyamus muticus hairy roots.fast-growing hairy root cultures of hyoscyamus muticus induced by agrobacterium rhizogenes offer a potential production system for tropane alkaloids. oxygen deficiency has been shown to limit growth and biomass accumulation of hairy roots, whereas little experimental data is available on the effect of oxygen on alkaloid production. we have investigated the effect of vitreoscilla hemoglobin (vhb) expression and cultivation conditions on the complete alkaloid profile of h. muticus hairy roots in s ...200616599546
expression of vitreoscilla hemoglobin enhances growth and levels of alpha-amylase in schwanniomyces occidentalis.a metabolic engineering approach was exploited to improve growth and protein secretion in the non-conventional yeast, schwanniomyces occidentalis. vitreoscilla hemoglobin (vhb) gene was expressed in s. occidentalis under the control of the native alpha-amylase (amy1) promoter. expression of vhb was confirmed by reverse transcriptase polymerase chain reaction and western blot hybridization analysis. effect of vhb on growth and protein secretion was studied in synthetic medium under both limiting ...200616642333
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