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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
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Journal of Microbiology and Biotechnology
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Journal DOI :
The Korean Society for Applied Microbiology and Biotechnology
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Volume & Issues
Volume 17, Issue 12 - Dec 2007
Volume 17, Issue 11 - Nov 2007
Volume 17, Issue 10 - Oct 2007
Volume 17, Issue 9 - Sep 2007
Volume 17, Issue 8 - Aug 2007
Volume 17, Issue 7 - Jul 2007
Volume 17, Issue 6 - Jun 2007
Volume 17, Issue 5 - May 2007
Volume 17, Issue 4 - Apr 2007
Volume 17, Issue 3 - Mar 2007
Volume 17, Issue 2 - Feb 2007
Volume 17, Issue 1 - Jan 2007
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Nanotechnology in Biodevices
Choi, Jeong-Woo ; Oh, Byung-Keun ; Kim, Young-Kee ; Min, Jun-Hong ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 5~14
Nanotechnology is the creation and utilization of materials, devices, and systems through the control of matter on the nanometer. The technology has been applied to biodevices such as bioelectronics and biochips to improve their performances. Nanoparticles, such as gold (Au) nanoparticles, are the most widely used of the various other nanotechnologies for manipulation at the nanoscale as well as nanobiosensors. The immobilization of biomolecules is playing an increasingly important role in the development of biodevices with high performance. Nanopatteming technology, which is able to increase the density of chip arrays, offers several advantages, including cost lowering, simultaneous multicomponent detection, and the efficiency increase of biochemical reactions. A microftuidic system incorporated with control of nanoliter of fluids is also one of the main applications of nanotechnologies. This can be widely utilized in the various fields because it can reduce detection time due to tiny amounts of fluids, increase signal-to-noise ratio by nanoparticles in channel, and detect multi-targets simultaneously in one chamber. This article reviews nanotechnologies such as the application of nanoparticles for the detection of biomolecules, the immobilization of biomolecules at nanoscale, nanopatterning technologies, and the microfluidic system for molecular diagnosis.
Identification and Molecular Characterization of Novel cry1-Type Toxin Genes from Bacillus thuringiensis K1 Isolated in Korea
Li Ming Shun ; Choi Jae-Young ; Roh Jong-Yul ; Shim Hee-Jin ; Kang Joong-Nam ; Kim Yang-Su ; Wang Yong ; Yu Zi Niu ; Jin Byung-Rae ; Je Yeon-Ho ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 15~20
To clone novel cry1-type genes from the Bacillus thuringiensis K1 isolate, about 2.4-kb-long PCR fragments were amplified with two primer sets of ATG1-F/N400-R and 1BeATG1-F/N400-R. Using PCR-RFLP, three novel cry1-type genes, cry1-1, cry1-7, and cry1-44, were obtained from B. thuringiensis K1 and the complete coding sequences of these novel genes were analyzed. The Cry1-1, Cry1-7, and Cry1-44 proteins showed maximum similarities of about 78.0%, 99.7%, and 91.0% with the Cry1Ha1, Cry1Be1, and Cry1Ac2 proteins, respectively. These novel cry1-type genes were expressed using a baculovirus expression vector system and their insecticidal activities were investigated. Whereas all three novel genes were toxic to Plutella xylostella larvae, only Cry1-1 showed insecticidal activity against Spodoptera exigua larvae.
Isolation and Characterization of Biopolymers Extracted from the Bark of Acanthopanax sessiliflorus and Their Anticomplement Activity
Jeong Sang-Chul ; Yang Byung-Keun ; Jeong Yong-Tae ; Rao Koyyalamudi Sundar ; Song Chi-Hyun ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 21~28
The crude biopolymer (AS-S1) and endobiopolymer (AS-S2) were isolated from the dry stem bark of Acanthopanax sessiliflorus and tested for anti complement activity. The two potent anticomplement biopolymers, AS-1 and AS-2-Fr.I, were isolated by the combination of ion-exchange chromatography and gel filtration methods from the endo-biopolymers (AS-S2). The anticomplement activity of AS-1 (MW 12 kDa) and AS-2-Fr.I (MW 180 kDa) were found to be 84.4% and 100.0%, respectively, at the concentration of
. Activated pathway of the complement system occurred in both classical and alternative pathways, as evidenced by crossed immunoelectrophoresis(CIEP), where a major pathway was detected to be the classical one. It was found that the anticomplement activities of the periodate oxidized were decreased significantly, but those of pronase digested biopolymers of AS-1 and AS-2-Fr.I were decreased very little. The AS-1 contained 2,4,6-tri-O-methyl-D-glucitol, 2,3,6-tri-O-methyl-D-galacitol, and 2,3,6-tri-O-methyl-D-galacitol, which indicated that AS-1 contained a
glucopyranosyl residue and a
galactosyl residue. AS-2-Fr.I contained mainly 2,4-di-O-methyl-D-mannitol and 2,3,4-tri-O-methyl-D-galacitol, which contained
linked mannosyl and
linked galactosyl residues.
Cloning, Characterization, and Expression of Xylanase A Gene from Paenibacillus sp. DG-22 in Escherichia coli
Lee, Tae-Hyeong ; Lim, Pyung-Ok ; Lee, Yong-Eok ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 29~36
The xynA gene encoding the xylanase A of Paenibacillus sp. DG-22 was isolated with a DNA probe obtained by PCR amplification, using degenerated primers deduced from the amino acid residues of the known N-terminal region of the purified enzyme and the conserved region in the family 11 xylanases. The positive clones were screened on the LB agar plates supplemented with xylan, by the Congo-red staining method. The xynA gene consists of a 630-bp open reading frame encoding a protein of 210 amino acids, and the XynA preprotein contains a 28-residues signal peptide whose cleavage yields a l82-residues mature protein of a calculated molecular weight of 20,000Da and pI value of 8.77. The cloned DNA fragment also has another ORF of 873 nucleotides that showed 76% identity to the putative transcriptional activator of Bacillus halodurans C-125. Most of the xylanase activity was found in the periplasmic space of E. coli. The xynA gene was subcloned into pQE60 expression vector to fuse with six histidine-tag. The recombinant xylanase A was purified by heating and immobilized metal affinity chromatography. The optimum pH and temperature of the purified enzyme were 6.0 and
, respectively. This histidine-tagged xylanase A was less thermostable than the native enzyme.
Cloning, Expression, and Characterization of Bacillus sp. snu-7 Inulin Fructotransferase
Kim, Chung-Sei ; Hong, Chang-Ki ; Kim, Kyoung-Yun ; Wang, Xiu-Ling ; Kang, Su-Il ; Kim, Su-Il ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 37~43
A gene encoding inulin fructotransferase (di-D-fructofuranose 1,2': 2,3' dianhydride [DFA III]-producing IFTase, EC 18.104.22.168) from Bacillus sp. snu-7 was cloned. This gene was composed of a single, 1,353-bp open reading frame encoding a protein composed of a 40-amino acid signal peptide and a 410-amino acid mature protein. The deduced amino acid sequence was 98% identical to Arthrobacter globiformis C11-1 IFTase (DFA III-producing). The enzyme was successfully expressed in E. coli as a functionally active, His-tagged protein, and it was purified in a single step using immobilized metal affinity chromatography. The purified enzyme showed much higher specific activity (1,276 units/mg protein) than other DFA III-producing IFTases. The recombinant and native enzymes were optimally active in very similar pH and temperature conditions. With a 103-min half-life at
, the recombinant enzyme was as stable as the native enzyme. Acidic residues and cysteines potentially involved in the catalytic mechanism are proposed based on an alignment with other IFTases and a DFA IIIase.
Isolation and Characterization of Mucous Exopolysaccharide (EPS) Produced by Vibrio furnissii Strain VB0S3
Bramhachari P.V. ; Kishor P.B. Kavi ; Ramadevi R. ; Kumar Ranadheer ; Rao, B. Rama ; Dubey Santosh Kumar ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 44~51
Marine bacterial strains were isolated trom coastal regions of Goa and screened for the strains that produce the highest amount of mucous expolysaccharide (EPS). Our screening resulted in the identification of the strain Vibrio furnissii VB0S3 (hereafter called VB0S3), as it produced the highest EPS in batch cultures during the late logarithmic growth phase. The isolate was identified as VB0S3 based on morphological and biochemical properties. Growth and EPS production were studied in mineral salts medium supplemented with NaCl (1.5%) and glucose (0.2%). The exopolymer was recovered from the culture supernatant by using three volumes of cold ethanol precipitation and dialysis procedure. Chemical analyses of EPS revealed that it is primarily composed of neutral sugars, uronic acids, and proteins. Fourier-transform infrared (FT-IR) spectroscopy revealed the presence of carboxyl, hydroxyl, and amide groups, which correspond to a typical heteropolymeric polysaccharide, and the EPS also possessed good emulsification activity. The gas chromatographic analysis of an alditol-acetate derivatized sample of EPS revealed that it was mainly composed of galactose and glucose. Minor components found were mannose, rhamnose, fucose, ribose, arabinose, and xylose. EPS was readily isolated from culture supernatants, which suggests that the EPS was a slime-like exopolysaccharide. This is the first report of exopolysaccharide characterization that describes the isolation and characterization of an EPS expressed by Vibrio surnissii strain VB0S3. The results of the study contribute significantly and go a long way towards an understanding of the correlation between growth and EPS production, chemical composition, and industrial applications of the exopolysaccharide in environmental biotechnology and bioremediation.
Microcosm Study for Revegetation of Barren Land with Wild Plants by Some Plant Growth-Promoting Rhizobacteria
Ahn, Tae-Seok ; Ka, Jong-Ok ; Lee, Geon-Hyoung ; Song, Hong-Gyu ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 52~57
Growth promotion of wild plants by some plant growth-promoting rhizobacteria (PGPR) was examined in the microcosms composed of soils collected separately from a grass-covered site and a nongrass-covered site in a lakeside barren area at Lake Paro, Korea. After sowing the seeds of eight kinds of wild plants and inoculation of several strains of PGPR, the total bacterial number and microbial activity were measured during 5 months of study period, and the plant biomasses grown were compared at the end of the study. Acridine orange direct counts in the inoculated microcosms,
in the soil from the grass-covered area and
in the soil from the nongrass-covered site, were almost twice higher than those in the uninoculated microcosms. The number of Pseudomonas sp., well-known bacteria as PGPR, and the soil dehydrogenase activity were also higher in the inoculated soils than the uninoculated soils. The first germination of sowed seeds in the inoculated microcosm was 5 days earlier than the uninoculated microcosm. Average lengths of all plants grown during the study period were 26% and 29% longer in the inoculated microcosms starting with the grass-covered soil and the nongrass-covered soil, respectively, compared with those in the uninoculated microcosms. Dry weights of whole plants grown were 67-82% higher in the inoculated microcosms than the uninoculated microcosms. Microbial population and activity and growth promoting effect by PGPR were all higher in the soils collected from the grass-covered area than in the nongrass-covered area. The growth enhancement of wild plants seemed to occur by the activities of inoculated microorganisms, and this capability of PGPR may be utilized for rapid revegetation of some barren lands.
Cloning and Overexpression of a Paenibacillus
in Pichia pastoris: Purification and Characterization of the Recombinant Enzyme
Yang, Peilong ; Shi, Pengjun ; Wang, Yaru ; Bai, Yingguo ; Meng, Kun ; Luo, Huiying ; Yuan, Tiezheng ; Yao, Bin ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 58~66
Isolation, expression, and characterization of a novel
with high specific activity and homology to Bacillus lichenases is described. One clone was screened from a genomic library of Paenibacillus sp. F-40, using lichenan-containing plates. The nucleotide sequence of the clone contains an ORF consisting of 717 nucleotides, encoding a
protein of 238 amino acids and 26 residues of a putative signal peptide at its N-terminus. The amino acid sequence showed the highest similarity of 87% to other
of Bacillus. The gene fragment Bg1 containing the mature glucanase protein was expressed in Pichia pastoris at high expression level in a 3-1 high-cell-density fermenter. The purified recombinant enzyme Bg1 showed activity against barley
, lichenan, and laminarin. The gene encodes an
(E. C. 22.214.171.124). When lichenan was used as substrate, the optimal pH was 6.5, and the optimal temperature was
values for lichenan are 2.96mg/ml,
, respectively. For barley
the values are 3.73mg/ml,
, respectively. The recombinant Bg1 had resistance to pepsin and trypsin. Other features of recombinant Bg1 including temperature and pH stability, and sensitivity to some metal ions and chemical reagents were also characterized.
Monitoring of Microbial Diversity and Activity During Bioremediation of Crude Oil-Contaminated Soil with Different Treatments
Baek, Kyung-Hwa ; Yoon, Byung-Dae ; Kim, Byung-Hyuk ; Cho, Dae-Hyun ; Lee, In-Sook ; Oh, Hee-Mock ; Kim, Hee-Sik ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 67~73
The present study compared the microbial diversity and activity during the application of various bioremediation processes to crude oil-contaminated soil. Five different treatments, including natural attenuation (NA), biostimulation (BS), biosurfactant addition (BE), bioaugmentation (BA), and a combined treatment (CT) of biostimulation, biosurfactant addition, and bioaugmentation, were used to analyze the degradation rate and microbial communities. After 120 days, the level of remaining hydrocarbons after all the treatments was similar, however, the highest rate (k) of total petroleum hydrocarbon (TPH) degradation was observed with the CT treatment (P<0.05). The total bacterial counts increased during the first 2 weeks with all the treatments, and then remained stable. The bacterial communities and alkane monooxygenase gene fragment, alkB, were compared by denaturing gradient gel electrophoresis (DGGE). The DGGE analyses of the BA and CT treatments, which included Nocardia sp. H17-1, revealed a simple dominant population structure, compared with the other treatments. The Shannon-Weaver diversity index (H') and Simpson dominance index (D), calculated from the DGGE profiles using 16S rDNA, showed considerable qualitative differences in the community structure before and after the bioremediation treatment as well as between treatment conditions.
Molecular Cloning and Functional Expression of esf Gene Encoding Enantioselective Lipase from Serratia marcescens ES-2 for Kinetic Resolution of Optically Active (S)-Flurbiprofen
Lee, Kwang-Woo ; Bae, Hyun-Ae ; Lee, Yong-Hyun ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 74~80
An enantioselective lipase gene (esf) for the kinetic resolution of optically active (S)-flurbiprofen was cloned from the new strain Serratia marcescens ES-2. The esf gene was composed of a 1,845-bp open reading frame encoding 614 amino acid residues with a calculated molecular mass of 64,978 Da. The lipase expressed in E. coli was purified by a three-step procedure, and it showed preferential substrate specificity toward the medium-chain-length fatty acids. The esf gene encoding the enantioselective lipase was reintroduced into the parent strain S. marcescens ES-2 for secretory overexpression. The transformant S. marcescens BESF secreted up to 217kU/ml of the enantioselective lipase, about 54-fold more than the parent strain, after supplementing 3.0% Triton X-207. The kinetic resolution of (S)-flurbiprofen was carried out even at an extremely high (R,S)-flurbiprofen ethyl ester [(R,S)-FEE] concentration of 500 mM, 130 kU of the S. marcescens ES-2 lipase per mmol of (R,S)-FEE, and 1,000 mM of succinyl
as the dispenser at
for 12h, achieving the high enantiomeric excess and conversion yield of 98% and 48%, respectively.
Distinct Regulation of the sprC Gene Encoding Streptomyces griseus Protease C from Other Chymotrypsin Genes in Streptomyces griseus IFO13350
Choi, Eun-Yong ; Oh, Eun-A ; Kim, Jong-Hee ; Kang, Dae-Kyung ; Hong, Soon-Kwang ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 81~88
The sprC gene encodes Streptomyces griseus protease C (SGPC), a bacterial chymotrypsin-like serine protease. Because the published data on sprC was not complete, we cloned and analyzed a new DNA fragment spanning downstream to upstream of the sprC gene from S. griseus IFO13350. The cloned 2.3-kb DNA fragment was placed on a high-copy number plasmid and introduced into Streptomyces lividans TK24. Chymotrypsin activity of the transformant was 8.5 times higher than that of the control after 3 days of cultivation and stably maintained until 9 days of cultivation, which dearly indicated that the cloned 2.3-kb fragment contained the entire sprC gene with its own promoter. When the same construct was introduced in the S. griseus IFO13350 (wild strain) and its two mutant strains in the A-factor regulatory cascade,
and HO1, the chymotrypsin activity increased fivefold only in the
strain. Transcriptional analysis based on RT-PCR revealed that the sprC gene is normally transcribed in both strains; however, earlier transcription was observed in the wild strain compared with the
strain. A gel mobility shift assay showed that the AdpA protein did not bind to the promoter region of sprC. All these data clearly indicate that the expression of sprC is not dependent on the AdpA protein, but is distinctly regulated from other chymotrypsin genes composing an AdpA regulon. Earlier morphological differentiation was observed in S. lividans TK24, and S. griseus IFO13350 and HO1, transformed with the expression vector. The transformant of S. griseus
formed markedly larger colonies. Antisense repression of sprC resulted in severe decrease of chymotrypsin activity, down to one-third of the control, and delayed morphological differentiation. All these data suggest that SGPC is related to normal morphogenesis in S. griseus.
Enhanced Proteomic Analysis of Streptomyces peucetius Cytosolic Protein Using Optimized Protein Solubilization Protocol
Lee, Kwang-Won ; Song, Eun-Jung ; Kim, June-Hyung ; Lee, Hei-Chan ; Liou, Kwang-Kyoung ; Sohng, Jae-Kyung ; Kim, Byung-Gee ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 89~95
Improvements in the dissolution of proteins in two-dimensional gel electrophoresis have greatly advanced the ability to analyze the proteomes of microorganisms under a wide variety of physiological conditions. This study examined the effect of various combinations of chaotropic agents, a reducing agent, and a detergent on the dissolution of the Streptomyces peucetius cytosolic proteins. The use of urea alone in a rehydration buffer as a chaotropic agent gave the proteome a higher solubility than any of the urea and thiourea combinations, and produced the highest resolution and clearest background in two-dimensional gel electrophoresis. Two % CHAPS, as a detergent in a rehydration buffer, improved the protein solubility. After examining the effect of several concentrations of reducing agent, 50 mM DTT in a rehydration buffer was found to be an optimal condition for the proteome analysis of Streptomyces. Using this optimized buffer condition, more than 2,000 distinct and differentially expressed soluble proteins could be resolved using two-dimensional gel electrophoresis with a pI ranging from 4-7. Under this optimized condition, 15 novel small proteins with low-level expression, which could not be analyzed under the non-optimized conditions, were identified. Overall, the optimized condition helped produce a better reference gel for Streptomyces peucetius.
Two Bacterial Entophytes Eliciting Both Plant Growth Promotion and Plant Defense on Pepper (Capsicum annuum L.)
Kang, Seung-Hoon ; Cho, Hyun-Soo ; Cheong, Hoon ; Ryu Choong-Min ; Kim, Ji-Hyun ; Park, Seung-Hwan ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 96~103
Plant growth-promoting rhizobacteria (PGPR) have the potential to be used as microbial inoculants to reduce disease incidence and severity and to increase crop yield. Some of the PGPR have been reported to be able to enter plant tissues and establish endophytic populations. Here, we demonstrated an approach to screen bacterial endophytes that have the capacity to promote the growth of pepper seedlings and protect pepper plants against a bacterial pathogen. Initially, out of 150 bacterial isolates collected from healthy stems of peppers cultivated in the Chungcheong and Gyeongsang provinces of Korea, 23 putative endophytic isolates that were considered to be predominating and representative of each pepper sample were selected. By phenotypic characterization and partial 16S rDNA sequence analysis, the isolates were identified as species of Ochrobacterium, Pantoea, Pseudomonas, Sphingomonas, Janthinobacterium, Ralstonia, Arthrobacter, Clavibacter, Sporosarcina, Acidovorax, and Brevundimonas. Among them, two isolates, PS4 and PS27, were selected because they showed consistent colonizing capacity in pepper stems at the levels of
tissue, and were found to be most closely related to Pseudomonas rhodesiae and Pantoea ananatis, respectively, by additional analyses of their entire 16S rDNA sequences. Drenching application of the two strains on the pepper seedlings promoted significant growth of peppers, enhancing their root fresh weight by 73.9% and 41.5%, respectively. The two strains also elicited induced systemic resistance of plants against Xanthomonas axonopodis pv. vesicatoria.
Acid (GABA) by Lactobacillus buchneri Isolated from Kimchi and its Neuroprotective Effect on Neuronal Cells
Cho, Yu-Ran ; Chang, Ji-Yoon ; Chang, Hae-Choon ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 104~109
Lactic acid bacteria that accumulated
acid (GABA) in culture medium were screened to identify strains with high GAB A-producing ability. One strain, MS, which was isolated from kimchi, showed the highest GABA-producing ability among the screened strains. MS was identified as Lactobacillus buchneri based on Gram-staining, metabolic characteristics, and 16S rDNA sequence determination, Optimum culture conditions for GABA production were determined: MRS broth containing 5% MSG, 1% NaCl, and 1% glucose, at an initial pH of 5.0, the incubation temperature at
for 36 h. Under these conditions, MS produced GABA at a concentration of 251 mM with a 94% GABA conversion rate. Moreover, culture extracts of Lb. buchneri MS partially or completely protected neuronal cells against neurotoxicantinduced cell death.
Enrichment of Electrochemically Active Bacteria Using a Three-Electrode Electrochemical Cell
Yoon, Seok-Min ; Choi, Chang-Ho ; Kim, Mi-A ; Hyun, Moon-Sik ; Shin, Sung-Hye ; Yi, Dong-Heui ; Kim, Hyung-Joon ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 110~115
Electrochemically active bacteria were successfully enriched in an electrochemical cell using a positively poised working electrode. The positively poised working electrode (+0.7 V vs. Ag/AgCl) was used as an electron acceptor for enrichment and growth of electrochemically active bacteria. When activated sludge and synthetic wastewater were fed to the electrochemical cell, a gradual increase in amperometric current was observed. After a period of time in which the amperometric current was stabilized (generally 8 days), linear correlations between the amperometric signals from the electrochemical cell and added BOD (biochemical oxygen demand) concentrations were established. Cyclic voltammetry of the enriched electrode also showed prominent electrochemical activity. When the enriched electrodes were examined with electron microscopy and confocal scanning laser microscopy, a biofilm on the enriched electrode surface and bacterium-like particles were observed. These experimental results indicate that the electrochemical system in this study is a useful tool for the enrichment of an electrochemically active bacterial consortium and could be used as a novel microbial biosensor.
Development of Bioreactor System for L-Tyrosine Synthesis Using Thermostable Tyrosine Phenol-Lyase
Kim, Do-Young ; Rha, Eugene ; Choi, Su-Lim ; Song, Jae-Jun ; Hong, Seung-Pyo ; Sung, Moon-Hee ; Lee, Seung-Goo ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 116~122
An efficient enzyme system for the synthesis of L-tyrosine was developed using a fed-batch reactor with continuous feeding of phenol, pyruvate, and ammonia. A thermo- and chemostable tyrosine phenol-lyase from Symbiobacterium toebii was employed as the biocatalyst in this work. The enzyme was produced using a constitutive expression system in Escherichia coli BL21, and prepared as a soluble extract by rapid clarification, involving treatment with 40% methanol in the presence of excess ammonium chloride. The stability of the enzyme was maintained for at least 18 h under the synthesis conditions, including 75 mM phenol at pH 8.5 and
. The fed-batch system (working volume, 0.51) containing 1.0 kU of the enzyme preparation was continuously fed with two substrate preparations: one containing 2.2 M phenol and 2.4 M sodium pyruvate, and the other containing 0.4 mM pyridoxal-5-phosphate and 4M ammonium chloride (pH 8.5). The system produced 130g/I of L-tyrosine within 30h, mostly as precipitated particles, upon continuous feeding of the substrates for 22 h. The maximum conversion yield of L-tyrosine was 94% on the basis of the supplied phenol.
Molecular Cloning and Characterization of Trehalose Biosynthesis Genes from Hyperthermophilic Archaebacterium Metallosphaera hakonesis
Seo, Ju-Seok ; An, Ju-Hee ; Baik, Moo-Yeol ; Park, Cheon-Seok ; Cheong, Jong-Joo ; Moon, Tae-Wha ; Park, Kwan-Hwa ; Choi, Yang-Do ; Kim, Chung-Ho ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 123~129
biosynthesis genes MhMTS and MhMTH, encoding a maltooligosyltrehalose synthase (MhMTS) and a maltooligosyltrehalose trehalohydrolase (MhMTH), respectively, have been cloned from the hyperthermophilic archaebacterium Metallosphaera hakonesis. The ORF of MhMTS is 2,142 bp long, and encodes 713 amino acid residues constituting a 83.8 kDa protein. MhMTH is 1,677 bp long, and encodes 558 amino acid residues constituting a 63.7 kDa protein. The deduced amino acid sequences of MhMTS and MhMTH contain four regions highly conserved for MTSs and three for MTHs that are known to constitute substrate-binding sites of starch-hydrolyzing enzymes. Recombinant proteins obtained by expressing the MhMTS and MhMTH genes in E. coli catalyzed a sequential reaction converting maltooligosaccharides to produce trehalose. Optimum pH of the MhMTS/MhMTH enzyme reaction was around 5.0 and optimum temperature was around 70 C. Trehalose-producing activity of the MhMTS/ MhMTH was notably stable, retaining 80% of the activity after preincubation of the enzyme mixture at
for 48 h, but was gradually abolished by incubating at above
. Addition of thermostable
increased the yield of trehalose production from maltopentaose by 10%. The substrate specificity of the MhMTS/MhMTH-catalyzed reaction was extended to soluble starch, the most abundant maltodextrin in nature.
Ectopic Expression of Apple MbR7 Gene Induced Enhanced Resistance to Transgenic Arabidopsis Plant Against a Virulent Pathogen
Lee, Soo-Yeon ; Choi, Yeon-Ju ; Ha, Young-Mie ; Lee, Dong-Hee ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 130~137
A disease resistance related gene, MbR7, was identified in the wild apple species, Malus baccata. The MbR7 gene has a single open reading frame (ORF) of 3,288 nucleotides potentially encoding a 1,095-amino acid protein. Its deduced amino acid sequence resembles the N protein of tobacco and the NL27 gene of potato and has several motifs characteristic of a TIR-NBS-LRR R gene subclass. Ectopic expression of MbR7 in Arabidopsis enhanced the resistance against a virulent pathogen, Pseudomonas syringae pv. tomato DC3000. Microarray analysis confirmed the induction of defense-related gene expression in 35S::MbR7 heterologous Arabidopsis plants, indicating that the MbR7 gene likely activates a downstream resistance pathway without interaction with pathogens. Our results suggest that MbR7 can be a potential target gene in developing a new disease-resistant apple variety.
Substitution of Pro206 and Ser86 Residues in the Retinal Binding Pocket of Anabaena Sensory Rhodopsin is Not Sufficient for Proton Pumping Function
Choi, Ah-Reum ; Kim, So-Young ; Yoon, Sa-Ryong ; Bae, Ki-Ho ; Jung, Kwang-Hwang ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 138~145
Anabaena sensory rhodopsin is a seven transmembrane protein that uses all-trans/13-cis retinal as a chromophore. About 22 residues in the retinal-binding pocket of microbial rhodopsins are conserved and important to control the quality of absorbing light and the function of ion transport or sensory transduction. The absorption maximum is 550 nm in the presence of all-trans retinal at dark. Here, we mutated Pro206 to Glu or Asp, of which the residue is conserved as Asp among all other microbial rhodopsins, and the absorption maximum and pKa of the proton acceptor group were measured by absorption spectroscopy at various pHs. Anabaena rhodopsin was expressed best in Escherichia coli in the absence of extra leader sequence when exogenous all-trans retinal was added. The wild-type Anabaena rhodopsin showed small absorption maximum changes between pH4 and 11. In addition, Pro206Asp showed 46 nm blue-shift at pH7.0. Pro206Glu or Asp may change the contribution to the electron distribution of the retinal that is involved in the major role of color tuning for this pigment. The critical residue Ser86 (Asp 96 position in bacteriorhodopsin: proton donor) for the pumping activity was replaced with Asp, but it did not change the proton pumping activity of Anabaena rhodopsin.
The Within-Host Population Dynamics of Normal Flora in the Presence of an Invading Pathogen and Antibiotic Treatments
Kim, Jung-Mo ; Lee, Dong-Hwan ; Song, Yoon-Seok ; Kang, Seong-Woo ; Kim, Seung-Wook ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 146~153
A mathematical competition model between normal flora and an invading pathogen was devised to allow analysis of bacterial infections in a host. The normal flora includes the various microorganisms that live on or within the host and act as a primary human immune system. Despite the important role of the normal flora, no mathematical study has been undertaken on models of the interaction between it and invading pathogens against a background of antibiotic treatment. To quantify key elements of bacterial behavior in a host, pairs of nonlinear differential equations were used to describe three categories of human health conditions, namely, healthy, latent infection, and active infection. In addition, a cutoff value was proposed to represent the minimum population level required for survival. The recovery of normal flora after antibiotic treatment was also included in the simulation because of its relation to human health recovery. The significance of each simulation parameter for the bacterial growth model was investigated. The devised simulation showed that bacterial proliferation rate, carrying capacity, initial population levels, and competition intensity have a significant effect on bacterial behavior. Consequently, a model was established to describe competition between normal flora and an infiltrating pathogen. Unlike other population models, the recovery process described by the devised model can describe the human health recovery mechanism.
Expression Profiles and Pathway Analysis in HEK 293 T Cells Overexpressing HIV-1 Tat and Nucleocapsid Using cDNA Microarray
Park, Seong-Eun ; Lee, Min-Joo ; Yang, Moon-Hee ; Ahn, Ka-Young ; Jang, Soo-In ; Suh, Young-Ju ; Myung, Hee-Joon ; You, Ji-Chang ; Park, Jong-Hoon ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 154~161
Human immunodeficiency virus type 1 (HIV-1) infections are responsible for a substantial number of deaths annually and represent a significant threat to public health. According to the latest study, the Tat (Transactivator of transcription) protein is essential in transcription and replication of viral genes, and is among the early expression genes involved in the life cycle of HIV. The virion NC (nucleocapsid) plays an important role in early mRNA expression and contributes to the rapid viral replication that occurs during HIV-1 infection. Therefore, we attempted to elucidate the relationship between the Tat protein and nucleocapsid protein. In a comparison of two independently prepared and hybridized samples, flag NC overexpressed HEK 293T cells and pTat overexpressed HEK 293T cells, and hybridization showed the differences in expression in each case. Among the microarray results confirmed with real-time reverse transcriptase assay, twelve genes were identified to be involved according to their gene expression profiles. Of approximately 8,208 human genes that were analyzed, we monitored candidate genes that might have been related to NC and Tat genes from gene expression profiles. Additionally, the pathways could be viewed and analyzed through the use of Pathway Studio software. The pathways from the gene list were built and paths were found among the molecules/cell objects/processes by the curation method.
Construction of Conjugative Gene Transfer System Between E. coli and Moderately Thermophilic, Extremely Acidophilic Acidithiobacillus caldus MTH-04
Liu, Xianggmei ; Lin, Jianqun ; Zhang, Zheng ; Bian, Jiang ; Zhao, Qing ; Liu, Ying ; Lin, Jianqiang ; Yan, Wangming ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 162~167
A genetic transfer system for introducing foreign genes to biomining microorganisms is urgently needed. Thus, a conjugative gene transfer system was investigated for a moderately thermophilic, extremely acidophilic biomining bacterium, Acidithiobacillus caldus MTH-04. The broad-hostrange IncP plasmids RP4 and R68.45 were transferred directly into A. caldus MTH-04 from Escherichia coli by conjugation at relatively high frequencies. Additionally the broad-hostrange IncQ plasmids pJRD215, pVLT33, and pVLT35 were also transferred into A. caldus MTH-04 with the help of plasmid RP4 or strains with plasmid RP4 integrated into their chromosome, such as E. coli SM10. The
selectable markers from these plasmids were successfully expressed in A. caldus MTH-04. Futhermore, the IncP and IncQ plasmids were transferred back into E. coli cells from A. caldus MTH-04, thereby confirming the initial transfer of these plasmids from E. coli to A. caldus MTH-04. All the IncP and IncQ plasmids studied were stable in A. caldus MTH-04. Consequently, this development of a conjugational system for A. caldus MTH-04 will greatly facilitate its genetic study.
'Bring to Lab' of 19 Novel Species Among 60 Isolates Retrieved from a Freshwater Pond
Song, Jae-Ho ; Yang, Seung-Jo ; Cho, Jang-Cheon ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 168~175
We report here on the cultivation of numerous novel bacterial species from a eutrophic freshwater pond. A total of 60 strains, 15 strains per each culture medium, were obtained from the surface of a eutrophic freshwater pond by employing a conventional dilution-plating method with four different kinds of culture media, including R2A, 1/10R2A, PCA, and 1/10PCA. Among the 60 strains isolated, 27 strains showed less than 97% 16S rRNA gene sequence similarities to validly published species, and thus they are considered to comprise 19 novel species. Of the 27 strains assigned to the novel species, the majority of the strains (20 strains) were affiliated with the Alphaproteobacteria and Betaproteobacteria. The remaining 7 strains were affiliated with the Gammaproteobacteria, Firmicutes, Actinobacteria, and Deinococci. Because we have isolated 19 novel species from a usual freshwater pond using a conventional culturing technique, our results suggest that an unexplored ecosystem, even if it looks like a common ecosystem found elsewhere, harbors diverse unidentified microbes, which will be definitely further characterized.
Cadaverine Protects Vibrio vulnificus from Superoxide Stress
Kang, In-Hye ; Kim, Ju-Sim ; Kim, Eui-Jin ; Lee, Jeong ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 176~179
An electron paramagnetic resonance (EPR) signal characteristic of the 5,5'-dimethyl-l-pyrroline-N-oxide (DMPO)-OH spin adduct, which is formed from the reaction of DMPO with superoxide radicals generated by xanthine oxidasemediated reaction, was significantly reduced by the cadaverine or Escherichia coli Mn-containing superoxide dismutase (MnSOD). Likewise, cytochrome c reduction by superoxide was inhibited by cadaverine, and the inhibition level increased in proportion to the level of cadaverine. The cadA mutant of Vibrio vulnificus, which does not produce cadaverine because of the lack of lysine decarboxylase, exhibits less tolerance to superoxide stress in comparison with wild type. The results indicate that cadaverine scavenges superoxide radicals, and protects cells from oxidative stress.
R-Type Pyocin is Required for Competitive Growth Advantage Between Pseudomonas aeruginosa Strains
Heo Yun-Jeong ; Chung In-Young ; Choi, Kelly B. ; Cho, You-Hee ;
Journal of Microbiology and Biotechnology, volume 17, issue 1, 2007, Pages 180~185
R-type pyocin is a bacteriophage tail-shaped bacteriocin produced by Pseudomonas aeruginosa, but its physiological roles are relatively unknown. Here we describe a role of R-type pyocin in the competitive growth advantages between P aeruginosa strains. Partial purification and gene disruption revealed that the major killing activity from the culture supernatant of PA14 is attributed to R-type pyocin, neither F-type nor S-type pyocins. These findings may provide insight into the forces governing P aeruginosa population dynamics to promote and maintain its biodiversity.