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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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Chitinophaga soli sp. nov. and Chitinophaga terrae sp. nov., Isolated from Soil of a Ginseng Field in Pocheon Province, Korea
An, Dong-Shan ; Im, Wan-Taek ; Lee, Sung-Taik ; Choi, Woo-Young ; Yoon, Min-Ho ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 705~711
Two novel strains of the Cytophaga-Flexibacter-Bacteroides(CFB) group, designated Gsoil
, were isolated from soil of a ginseng field of Pocheon Province in Korea. Both strains were Gram-negative, aerobic, nonmotile, nonspore-forming, and rod-shaped. Phylogenetic analysis based on 16S rRNA gene sequences indicated that both isolates belong to the genus Chitinophaga but were clearly separated from established species of this genus. The sequence similarities between strain Gsoil
and type strains of the established species and between strain Gsoil
and type strains of the established species ranged from 91.4 to 94.7% and 91.6 to 94.2%, respectively. Phenotypic and chemotaxonomic data(major menaquinone, MK-7; major fatty acids,
; major hydroxy fatty acid,
; major polyamine, homospermidine) supported the affiliation of both strains Gsoil
to the genus Chitinophaga. Furthermore, the results of physiological and biochemical tests allowed genotypic and phenotypic differentiation of both strains from the other validated Chitinophaga species. Therefore, the two isolates represent two novel species, for which the name Chitinophaga soli sp. nov.(type strain, Gsoil
) and Chitinophaga terrae sp. nov.(type strain, Gsoil
) are proposed.
Differences in Optimal pH and Temperature for Cell Growth and Antibody Production Between Two Chinese Hamster Ovary Clones Derived from the Same Parental Clone
Kim, Sung-Hyun ; Lee, Gyun-Min ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 712~720
To investigate clonal variations of recombinant Chinese hamster ovary(rCHO) clones in response to culture pH and temperature, serum-free suspension cultures of two antibody-producing CHO clones(clones A and B), which were isolated from the same parental clone by the limiting dilution method, were performed in a bioreactor at pH values in the range of 6.8-7.6, and two different temperatures,
. In regard to cell growth, clone A and clone B displayed similar responses to temperature, although their degree of response differed. In contrast, clones A and B displayed different responses to temperature in regard to antibody production. In the case of clone A, no significant increase in maximum antibody concentration was achieved by lowering the culture temperature. The maximum antibody concentration obtained at
(pH 7.4) and
(pH 7.0) were
, respectively. On the other hand, in the case of clone B, an approximately 2.5-fold increase in maximum antibody concentration was achieved by lowering the culture temperature. The enhanced maximum antibody concentration of clone B at
at pH 7.2) was due to not only enhanced specific antibody productivity but also to prolonged culture longevity. At
, the culture longevity of clone A also improved, but not as much as that of clone B. Taken together, CHO clones derived from the same parental clone displayed quite different responses to culture temperature and pH with regards antibody production, suggesting that environmental parameters such as temperature and pH should be optimized for each CHO clone.
Synthesis of L-threo-3,4-Dihydroxyphenylserine(L-threo-DOPS) with Thermostabilized Low-Specific L-Threonine Aldolase from Streptomyces coelicolor A3(2)
Baik, Sang-Ho ; Yoshioka, Hideki ; Yukawa, Hideaki ; Harayama, Shigeaki ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 721~727
Stability-enhanced mutants, H44, 11-94, 5A2-84, and F8, of L-threonine aldolase(L-TA) from Streptomyces coelicolor A3(2)(SCO1085) were isolated by an error-prone PCR followed by a high-throughput screening. Each of these mutant, had a single amino acid substitution: H177Y in the H44 mutant, A169T in the 11-94 mutant, D104N in the 5A2-84 mutant and F18I in the F8 mutant. The residual L-TA activity of the wild-type L-TA after a heat treatment for 20 min at
was only 10.6%. However, those in the stability-enhanced mutants were 85.7% for the H44 mutant, 58.6% for the F8 mutant, 62.1% for the 5A2-84 mutant, and 67.6% for the 11-94 mutant. Although the half-life of the wild-type L-TA at
was 1.3 min, those of the mutant L-TAs were longer: 14.6 min for the H44 mutant, 3.7 min for the 11-94 mutant, 5.8 min for the 5A2-84 mutant, and 5.0 min for the F8 mutant. The specific activity did not change in most of the mutants, but it was decreased by 45% in the case of mutant F8. When the aldol condensation of glycine and 3,4-dihydroxybenzaldehyde was studied by using whole cells of Escherichia coli containing the wild-type L-TA gene, L-threo-3,4-dihydroxyphenylserine(L-threo-DOPS) was successfully synthesized with a yield of 2.0 mg/ml after 20 repeated batch reactions for 100 h. However, the L-threo-DOPS synthesizing activity of the enzyme decreased with increased cycles of the batch reactions. Compared with the wild-type L-TA, H44 L-TA kept its L-threo-DOPS synthesizing activity almost constant during the 20 repeated batch reactions for 100 h, yielding 4.0 mg/ml of L-threo-DOPS. This result showed that H44 L-TA is more effective than the wild-type L-TA for the mass production of L-threo-DOPS.
pT7MT, a Metallothionein 2A-Tagged Novel Prokaryotic Fusion Expression Vector
Marikar, Faiz M.M.T. ; Fang, Lei ; Jiang, Shu-Han ; Hua, Zi-Chun ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 728~732
In the present article, a novel fusion expression vector for Escherichia coli was developed based on the pTORG plasmid, a derivative of pET32a. This vector, named pT7MT(GenBank Accession No DQ504436), carries a T7 promoter and it drives the downstream gene encoding Metallothionein 2A(MT2A). There are in-framed multiple cloning sites(MCS) downstream of the MT2A gene. A target gene can be cloned into the MCS and fused to the C-terminal of the MT2A gene in a compatible open reading frame(ORF) to achieve fusion expression. The metal-binding capability of MT2A allows the purification of fusion proteins by metal chelating affinity chromatography, known as
-affinity chromatography. Using this expression vector, we successfully got the stable and high-yield expression of MT2A-GST and MT2A-Troponin I fusion proteins. These two proteins were easily purified from the supernatant of cell lysates by one-step
-affinity chromatography. The final yields of MT2A-GST and MT2A-Troponin I were 30mg/l and 28mg/l in LB culture, respectively. Taken together, our data suggest that pT7MT can be applied as a useful expression vector for stable and high-yield production of fusion proteins.
Partial Purification and Characterization of Exoinulinase from Kluyveromyces marxianus YS-1 for Preparation of High-Fructose Syrup
Singh, Ram Sarup ; Dhaliwal, Rajesh ; Puri, Munish ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 733~738
An extracellular exoinulinase(
fructan fructanohydrolase, EC 220.127.116.11), which catalyzes the hydrolysis of inulin into fructose and glucose, was purified 23.5-fold by ethanol precipitation, followed by Sephadex G-100 gel permeation from a cell-free extract of Kluyveromyces marxianus YS-1. The partially purified enzyme exhibited considerable activity between pH 5 to 6, with an optimum pH of 5.5, while it remained stable(100%) for 3 h at the optimum temperature of
produced a 2A-fold and 1.2-fold enhancement in enzyme activity, whereas
completely inhibited the inulinase. A preparation of the partially purified enzyme effectively hydrolyzed inulin, sucrose, and raffinose, yet no activity was found with starch, lactose, and maltose. The enzyme preparation was then successfully used to hydrolyze pure inulin and raw inulin from Asparagus racemosus for the preparation of a high-fructose syrup. In a batch system, the exoinulinase hydrolyzed 84.8% of the pure inulin and 86.7% of the raw Asparagus racemosus inulin, where fructose represented 43.6mg/ml and 41.3mg/ml, respectively.
Production of Recombinant Polyhedra Containing Cry1Ac Fusion Protein in Insect Cell Lines
Kim, Jae-Su ; Choi, Jae-Young ; Roh, Jong-Yul ; Lee, Han-Young ; Jang, Seung-Sik ; Je, Yeon-Ho ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 739~744
Insect cell lines and the control of infection for obtaining the maximum amount of polyhedrin-Cry1Ac-polyhedrin fusion protein from Bactrus in monolayer and suspension culture systems were tested. Growth rates of the Trichoplusia ni(High-Five) cell line in both culture systems were better than the other insect cell lines, Spodoptera frugiferda(Sf-9, Sf-21), Trichoplusia ni(Tn5), and Spodoptera exigua(Se301). The expression of the fusion protein in a monolayer culture showed that Se301 cells were 2.3-4.8 times more productive on a per cell basis than the other cell lines. However, in suspension culture, only High-Five cells were productive. High-Five cells infected with Bactrus at a multiplicity of infection(MOI) of 5 and a cell density of
cells per ml were more productive than the other infection condition in a suspension culture suitable for a large-scale production of baculovirus. In conclusion, for the large-scale production of Bactrus in vitro, High-Five cells showing good growth and high productivity are suitable.
Purification and Characterization of Extracellular
-Glucosidase from Sinorhizobium kostiense AFK-13 and Its Algal Lytic Effect on Anabaena flos-aquae
Kim, Jeong-Dong ; Lee, Choul-Gyun ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 745~752
-glucosidase from the algal lytic bacterium Sinorhizobium kostiense AFK-13, grown in complex media containing cellobiose, was purified to homogeneity by successive ammonium sulfate precipitation, and anion-exchange and gel-filtration chromatographies. The enzyme was shown to be a monomeric protein with an apparent molecular mass of 52 kDa and isoelectric point of approximately 5.4. It was optimally active at pH 6.0 and
and possessed a specific activity of 260.4 U/mg of protein against
(pNPG). A temperature-stability analysis demonstrated that the enzyme was unstable at
and above. The enzyme did not require divalent cations for activity, and its activity was significantly suppressed by
, whereas sodium dodecyl sulfate(SDS) and Triton X-100 moderately inhibited the enzyme to under 70% of its initial activity. In an algal lytic activity analysis, the growth of cyanobacteria, such as Anabaena flos-aquae, A. cylindrica, A. macrospora, Oscillatoria sancta, and Microcystis aeruginosa, was strongly inhibited by a treatment of 20 ppm/disc or 30 ppm/disc concentration of the enzyme.
Diversity of Paenibacillus spp. in the Rhizosphere of Four Sorghum(Sorghum bicolor) Cultivars Sown with Two Contrasting Levels of Nitrogen Fertilizer Assessed by rpoB-Based PCR-DGGE and Sequencing Analysis
Coelho, Marcia Reed Rodrigues ; Mota, Fabio Faria Da ; Carneiro, Newton Portilho ; Marriel, Ivanildo Evodio ; Paiva, Edilson ; Rosado, Alexandre Soares ; Seldin, Lucy ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 753~760
The diversity of Paenibacillus species was assessed in the rhizospheres of four cultivars of sorghum sown in Cerrado soil amended with two levels of nitrogen fertilizer(12 and 120 kg/ha). Two cultivars(IS 5322-C and IS 6320) demanded the higher amount of nitrogen to grow, whereas the other two(FBS 8701-9 and IPA 1011) did not. Using the DNA extracted from the rhizospheres, a Paenibacillus-specific PCR system based on the RNA polymerase gene(rpoB) was chosen for the molecular analyses. The resulting PCR products were separated into community fingerprints by DGGE and the results showed a clear distinction between cultivars. In addition, clone libraries were generated from the rpoB fragments of two cultivars(IPA 1011 and IS 5322-C) using both fertilization conditions, and 318 selected clones were sequenced. Analyzed sequences were grouped into 14 Paenibacillus species. A greater diversity of Paenibacillus species was observed in cultivar IPA 1011 compared with cultivar IS 5322-C. Moreover, statistical analyses of the sequences showed that the bacterial diversity was more influenced by cultivar type than nitrogen fertilization, corroborating the DGGE results. Thus, the sorghum cultivar type was the overriding determinative factor that influenced the community structures of the Paenibacillus communities in the habitats investigated.
Biochemical and Genetic Characterization of Arazyme, an Extracellular Metalloprotease Produced from Serratia proteamaculans HY-3
Kwak, Jang-Yul ; Lee, Ki-Eun ; Shin, Dong-Ha ; Maeng, Jin-Soo ; Park, Doo-Sang ; Oh, Hyun-Woo ; Son, Kwang-Hee ; Bae, Kyung-Sook ; Park, Ho-Yong ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 761~768
Serratia proteamaculans HY-3 isolated from the digestive tract of a spider produces an extracellular protease named arazyme, with an estimated molecular mass of 51.5 kDa. The purified enzyme was characterized as having high activities at wide pH and temperature ranges. We further characterized biochemical features of the enzymatic reactions under various reaction conditions. The protease efficiently hydrolyzed a broad range of protein substrates including albumin, keratin, and collagen. The dependence of enzymatic activities on the presence of metal ions such as calcium and zinc indicated that the enzyme is a metalloprotease, together with the previous observation that the proteolytic activity of the enzyme was not inhibited by aspartate, cysteine, or serine protease inhibitors, but strongly inhibited by 1,10-phenanthroline and EDTA. The araA gene encoding the exoprotease was isolated as a 5.6 kb BamHI fragment after PCR amplification using degenerate primers and subsequent Southern hybridization. The nucleotide sequence revealed that the deduced amino acid sequences shared extensive similarity with those of the serralysin family of metalloproteases from other enteric bacteria. A gene(inh) encoding a putative protease inhibitor was also identified immediately adjacent to the araA structural gene.
Optimization of Ascorbic Acid-2-Phosphate Production from Ascorbic Acid Using Resting Cell of Brevundimonas diminuta
Shin, Woo-Jung ; Kim, Byung-Yong ; Bang, Won-Gi ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 769~773
With the aim to produce ascorbic acid-2-phosphate(AsA-2-P) from L-ascorbic acid(AsA, Vitamin C), nine bacteria conferring the ability to transform AsA to AsA-2-P were isolated from soil samples alongside known strains from culture collections. Most isolates were classified to the genus Brevundimonas by 16S phylogenetic analysis. Among them, Brevundimonas diminuta KACC 10306 was selected as the experimental strain because of its the highest productivity of AsA-2-P. The optimum set of conditions for the AsA-2-P production from AsA using resting cells as the source of the enzyme was also investigated. The optimum cultivation time was 16 h and the cell concentration was 120g/l(wet weight). The optimum concentrations of AsA and pyrophosphate were 550mM and 450mM, respectively. The most effective buffer was 50mM sodium formate. The optimum pH was 4.5 and temperature was
. Under the above conditions, 27.5g/l of AsA-2-P was produced from AsA after 36 h of incubation, which corresponded to a 19.7% conversion efficiency based on the initial concentration of AsA.
Characterization and Purification of Acidocin 1B, a Bacteriocin Produced by Lactobacillus acidophilus GP1B
Han, Kyoung-Sik ; Kim, Young-Hoon ; Kim, Sae-Hun ; Oh, Se-Jong ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 774~783
In the present study, acidocin 1B, a bacteriocin produced by Lactobacillus acidophilus GP 1B, exhibited profound inhibitory activity against a variety of LAB and pathogens, including Gram-negative bacteria, and its mode of action was to destabilize the cell wall, thereby resulting in bactericidal lysis. Acidocin 1B was found to be heat stable, because it lost no activity when it was heated up to
for 60 min. It retained approximately 67% of the initial activity after storage for 30 days at
, and 50% of its initial activity after 30 days at
. The molecular mass of acidocin 1B was estimated to be 4,214.65 Da by mass spectrometry. Plasmid curing results indicated that a plasmid, designated as pLA1B, seemed to be responsible for both acidocin 1B production and host immunity, and that the pLA1B could be transformed into competent cells of L. acidophilus ATCC 43121 by electroporation. Our findings indicate that the acidocin 1B and its producer strain may have potential value as a biopreservative in food systems.
Removal of Organic Load from Olive Washing Water by an Aerated Submerged Biofilter and Profiling of the Bacterial Community Involved in the Process
Pozo, Clementina ; Rodelas, Belen ; Martinez-Toledo, M. Victoria ; Vilchez, Ramiro ; Gonzalez-Lopez, Jesus ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 784~791
The present work aims to use a biofilter technology(aerated submerged filters) for the aerobic transformation at laboratory-scale of olive washing water(OWW) generated in the first steps of olive oil processing, as well as the genetic profiling and identification to the species level of the bacteria involved in the formation of the biofilm, by means of TGGE. Chemical parameters, such as biological oxygen demand at five days(
) and chemical oxygen demand(COD), decreased markedly(up to 90 and 85%, respectively) by the biological treatment, and the efficiency of the process was significantly affected by aeration and inlet flow rates. The total polyphenol content of inlet OWW was only moderately reduced(around 50% decrease of the inlet content) after the biofilter treatment, under the conditions tested. Partial 16S rRNA genes were amplified using total DNA extracted from the biofilm and separated by TGGE. Sequences of isolated bands were mostly affiliated to the
of Proteobacteria, and often branched in the periphery of bacteria] genera commonly present in soil(Rhizobium, Reichenowia, Agrobacterium, and Sphingomonas). The data obtained by the experimentation at laboratory scale provided results that support the suitability of the submerged filter technology for the treatment of olive washing waters with the purpose of its reutilization.
Cloning and Characterization of Glycogen-Debranching Enzyme from Hyperthermophilic Archaeon Sulfolobus shibatae
Van, Trinh Thi Kim ; Ryu, Soo-In ; Lee, Kyung-Ju ; Kim, Eun-Ju ; Lee, Soo-Bok ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 792~799
A gene encoding a putative glycogen-debranching enzyme in Sulfolobus shibatae(abbreviated as SSGDE) was cloned and expressed in Escherichia coli. The recombinant enzyme was purified to homogeneity by heat treatment and Ni-NTA affinity chromatography. The recombinant SSGDE was extremely thermostable, with an optimal temperature at
. The enzyme had an optimum pH of 5.5 and was highly stable from pH 4.5 to 6.5. The substrate specificity of SSGDE suggested that it possesses characteristics of both amylo-1,6-glucosidase and
-1,4-glucanotransferase. SSGDE clearly hydrolyzed pullulan to maltotriose, and
to maltose and
-cyclodextrin. At the same time, SSGDE transferred maltooligosyl residues to the maltooligosaccharides employed, and maltosyl residues to
. The enzyme preferentially hydrolyzed amylopectin, followed in a decreasing order by glycogen, pullulan, and amylose. Therefore, the present results suggest that the glycogen-debranching enzyme from S. shibatae may have industrial application for the efficient debranching and modification of starch to dextrins at a high temperature.
The Brown-Rot Basidiomycete Fomitopsis palustris Has the Endo-Glucanases Capable of Degrading Microcrystalline Cellulose
Yoon, Jeong-Jun ; Cha, Chang-Jun ; Kim, Yeong-Suk ; Son, Dong-Won ; Kim, Young-Kyoon ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 800~805
Two endoglucanases with processive cellulase activities, produced from Fomitopsis palustris grown on 2% microcrystalline cellulose(Avicel), were purified to homogeneity by anion-exchange and gel filtration column chromatography systems. SDS-PAGE analysis indicated that the molecular masses of the purified enzymes were 47 kDa and 35 kDa, respectively. The amino acid sequence analysis of the 47-kDa protein(EG47) showed a sequence similarity with fungal glycoside hydrolase family 5 endoglucanase from the white-rot fungus Phanerochaete chrysosporium. N-terminal and internal amino acid sequences of the 35-kDa protein(EG35), however, had no homology with any other glycosylhydrolases, although the enzyme had high specific activity against carboxymethyl cellulose, which is a typical substrate for endoglucanases. The initial rate of Avicel hydrolysis by EG35 was relatively fast for 48 h, and the amount of soluble reducing sugar released after 96 h was
. Although EG47 also hydrolyzed Avicel, the hydrolysis rate was lower than that of EG35. Thin layer chromatography analysis of the hydrolysis products released from Avicel indicated that the main product was cellobiose, suggesting that the brown-rot fungus possesses processive EGs capable of degrading crystalline cellulose.
Generation of a Specific Marker to Discriminate Bacillus anthracis from Other Bacteria of the Bacillus cereus Group
Kim, Hyoung-Tai ; Seo, Gwi-Moon ; Jung, Kyoung-Hwa ; Kim, Seong-Joo ; Kim, Jee-Cheon ; Oh, Kwang-Geun ; Koo, Bon-Sung ; Chai, Young-Gyu ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 806~811
Bacillus anthracis is a soil pathogen capable of causing anthrax that is closely related to several environmental species, including B. cereus, B. mycoides, and B. thuringiensis. DNA homology studies showed that B. anthracis, B. cereus, B. mycoides, and B. thuringiensis are closely related, with a high sequence homology. To establish a method to specifically detect B. anthracis in situations such as environmental contamination, we initially performed RAPD-PCR with a 10-mer random primer and confirmed the presence of specific PCR bands only in B. anthracis species. One region specific for B. anthracis was cloned and sequenced, and an internal primer set was designed to amplify a 241-bp DNA fragment within the sequenced region. The PCR system involving these specific primer sets has practical applications. Using lyses methods to prepare the samples for PCR, it was possible to quickly amplify the 241-bp DNA segment from samples containing only a few bacteria. Thus, the PCR detection method developed in this study is expected to facilitate the monitoring of environmental B. anthracis contamination.
Isolation and ars Detoxification of Arsenite-Oxidizing Bacteria from Abandoned Arsenic-Contaminated Mines
Chang, Jin-Soo ; Yoon, In-Ho ; Kim, Kyoung-Woong ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 812~821
The ecosystems of certain abandoned mines contain arsenic-resistant bacteria capable of performing detoxification when an ars gene is present in the bacterial genome. The ars gene has already been isolated from Pseudomonas putida and identified as a member of the membrane transport regulatory deoxyribonucleic acid family. The arsenite-oxidizing bacterial strains isolated in the present study were found to grow in the presence of 66.7 mM sodium arsenate(
), yet experienced inhibited growth when the sodium arsenite(
) concentration was higher than 26 mM. Batch experiment results showed that Pseudomonas putida strain OS-5 completely oxidized 1 mM of As(III) to As(V) within 35 h. An arsB gene encoding a membrane transport regulatory protein was observed in arsenite-oxidizing Pseudomonas putida strain OS-5, whereas arsB, arsH, and arrA were detected in strain OS-19, arsD and arsB were isolated from strain RW-18, and arsR, arsD, and arsB were found in E. coli strain OS-80. The leader gene of arsR, -arsD, was observed in a weak acid position. Thus, for bacteria exposed to weak acidity, the ars system may cause changes to the ecosystems of As-contaminated mines. Accordingly, the present results suggest that arsR, arsD, arsAB, arsA, arsB, arsC, arsH, arrA, arrB, aoxA, aoxB, aoxC, aoxD, aroA, and aroB may be useful for arsenite-oxidizing bacteria in abandoned arsenic-contaminated mines.
Cloning and Characterization of a Gene Encoding Phosphoketolase in a Lactobacillus paraplantarum Isolated from Kimchi
Jeong, Do-Won ; Lee, Jung-Min ; Lee, Hyong-Joo ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 822~829
A gene coding for phosphoketolase, a key enzyme of carbohydrate catabolism in heterofermentative lactic acid bacteria(LAB), was cloned from a Lactobacillus paraplantarum C7 and expressed in Escherichia coli. The gene is 2,502 bp long and codes for a 788-amino-acids polypeptide with a molecular mass of 88.7 kDa. A Shine-Dalgarno sequence(aaggag) and an inverted-repeat terminator sequence are located upstream and downstream of the phosphoketolase gene, respectively. The gene exhibits an identity of >52% with phosphoketolases of other LAB. The phosphoketolase of Lb. paraplantarum C7(LBPK) contains several highly conserved phosphoketolase signature regions and typical thiamine pyrophosphate(TPP) binding sites, as reported for other TPP-dependent enzymes. The phosphoketolase gene was fused to a glutathione S-transferase(GST::LBPK) gene for purification. The GST::LBPK fusion protein was detected in the soluble fraction of a recombinant Escherichia coli BL21. The GST::LBPK fusion protein was purified with a yield of 4.32mg/400ml by GSTrap HP affinity column chromatography and analyzed by N-terminal sequencing. LBPK was obtained by factor Xa treatment of fusion protein and the final yield was 3.78mg/400ml. LBPK was examined for its N-terminal sequence and phosphoketolase activity. The
values for fructose-6-phosphate were
, respectively, and the optimum temperature and pH for the production of acetyl phosphate were
and 7.0, respectively.
Cloning, Sequencing, and Characterization of the Pradimicin Biosynthetic Gene Cluster of Actinomadura hibisca P157-2
Kim, Byung-Chul ; Lee, Jung-Min ; Ahn, Jong-Seog ; Kim, Beom-Seok ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 830~839
Pradimicins are potent antifungal antibiotics having an unusual dihydrobenzo[
]naphthacenequinone aglycone substituted with D-alanine and sugars. Pradimicins are polyketide antibiotics produced by Actinomadura hibisca P157-2. The gene cluster involved in the biosynthesis of pradimicins was cloned and sequenced. The pradimicin gene cluster was localized to a 39-kb DNA segment and its involvement in the biosynthesis of pradimicin was proven by gene inactivation of prmA and prmB(ketosynthases
). The pradimicin gene cluster consists of 28 open reading frames(ORFs), encoding a type II polyketide synthase(PKS), the enzymes involved in sugar biosynthesis and tailoring enzymes as well as two resistance proteins. The deduced proteins showed strong similarities to the previously validated gene clusters of angucyclic polyketides such as rubromycin, griseorhodin, and fredericamycin. From the pradimicin gene cluster, prmP3 encoding a component of the acetyl-CoA carboxylase complex was disrupted. The production levels of pradimicins of the resulting mutants decreased to 62% of the level produced by the wild-type strain, which indicate that the acetyl-CoA carboxylase gene would have a significant role in the production of pradimicins through supplying the extender unit precursor, malonyl-CoA.
Expression and Purification of Recombinant Active Prostate-Specific Antigen from Escherichia coli
Jeong, Su-Jin ; Lee, Seong-Wook ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 840~846
Human prostate-specific antigen(PSA), a 33 kDa serine protease with comprehensive homology to glandular kallikrein, is secreted from prostatic tissue into the seminal fluid and enters into the circulation. The level of PSA increases in the serum of patients with prostatic cancer and hence is widely employed as a marker of the disease status. In particular, an enzymatically active PSA that is a form cleaved at the N-terminal seven-amino-acids prosequence, APLILSR, of proPSA may play an important roll in the progression of prostate cancer. Thus, the presence of the active form would selectively discriminate the cancer from benign prostatic hyperplasia. In this study, we developed a convenient purification method for the acquisition of active PSA and proPSA. Recombinant proPSA and active PSA were expressed directly in Escherichia coli, easily and efficiently isolated from inclusion bodies, refolded, and purified. Moreover, the enzymatic activity of the recombinant active PSA was confirmed as serine protease using chromogenic chymotrypsin substrate. This purified active PSA could be further applied to scrutinize the biological or conformational characteristics of the protein and to develop specific diagnostic and/or therapeutic agents against prostate cancer.
An Efficient Method for the Extraction of Astaxanthin from the Red Yeast Xanthophyllomyces dendrorhous
Choi, Seok-Keun ; Kim, Jeong-Hwan ; Park, Young-Sam ; Kim, Young-Jin ; Chang, Hyo-Ihl ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 847~852
This study investigated an efficient method for the extraction of astaxanthin from the red yeast Xanthophyllomyces dendrorhous. The extraction process comprised three steps: 1) cultivating the yeast; 2) treating the yeast culture suspension with microwaves to destroy the cell walls and microbodies; and 3) drying the yeast and extracting the astaxanthin pigment using ethanol, methanol, acetone, or a mixture of the three as the extraction solvent. Ultimately, various treatment tests were performed to determine the conditions for optimal pigment extraction, and the total carotenoid and astaxanthin contents were quantified. A frequency of 2,450 MHz, an output of 500 watts, and irradiation time of 60 s were the most optimum conditions for yeast cell wall destruction. Furthermore, optimal pigment extraction occurred when using a cell density of 10g/l at
over 24 h, with a 10% volume of ethanol.
Raceway Cultivation of Spirulina platensis Using Underground Water
Kim, Choong-Jae ; Jung, Yun-Ho ; Ko, So-Ra ; Kim, Hong-Ik ; Park, Yong-Ha ; Oh, Hee-Mock ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 853~857
The semi-outdoor cultivation of Spirulina platens is was attempted using an underground-water-based medium. Occurrence of contaminant organisms such as Chlorella sp. and Chlamydomonas sp. was not found from a microscopic observation and bacteria were not detected from denaturing gradient gel electrophoresis(DGGE) analysis of PCR-amplified 16S rDNA during the cultivation, owing to pH control and the high quality of the underground water. The mean productivity was high at
with a range of
despite the unfavorable weather conditions of the rainy season. The cultivated S. platens is included a normal protein content of 58.9%. Consequently, the underground water improved the biomass productivity and the biomass quality because of an abundant supplementation of natural minerals and through a contaminant-free culture.
Antibacterial Activity of Sophoraflavanone G Isolated from the Roots of Sophora flavescens
Cha, Jeong-Dan ; Jeong, Mi-Ran ; Jeong, Seung-Il ; Lee, Kyung-Yeol ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 858~864
This study investigated the antibacterial activities of sophoraflavanone G from Sophora flavescens in combination with two antimicrobial agents against oral bacteria. The combined effect of sophoraflavanone G and the antimicrobial agents was evaluated using the checkerboard method to obtain a fractional inhibitory concentration(FIC) index. The sophoraflavanone G+ampicillin(AM) combination was found to have a synergistic effect against S. mutans, S. sanguinis, S. sobrinus, S. gordonii, A. actinomycetemcomitans, F nucleatum, P. intermedia, and P. gingivalis, whereas the sophoraflavanone G+gentamicin(GM) combination had a synergistic effect against S. sanguinis, S. criceti, S. anginosus, A. actinomycetemcomitans, F nucleatum, P. intermedia, and P. gingivalis. Neither combination exhibited any antagonistic interactions(FIC index>4). In particular, the MICs/MBCs for all the bacteria were reduced to one-half
one-sixteenth as a result of the drug combinations. A synergistic interaction was also confirmed by time-kill studies for nine bacteria where the checkerboard suggested synergy. Thus, a strong bactericidal effect was exerted through the drug combinations, plus in vitro data suggested that sophoraflavanone G combined with other antibiotics may be microbiologically beneficial rather than antagonistic.
Redoxcitrinin, a Biogenetic Precursor of Citrinin from Marine Isolate of Fungus Penicillium sp.
Zhang, Dahai ; Li, Xianguo ; Kang, Jung-Sook ; Choi, Hong-Dae ; Jung, Jee-H. ; Son, Byeng-Wha ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 865~867
A chemical analysis of the fermentation of the marine-derived fungus Penicillium sp. led to the isolation of a biogenetic precursor of citrinin, redoxcitrinin(1), together with polyketide mycotoxins, phenol A(2), citrinin H2(3), 4-hydroxymellein(4), citrinin(5), and phenol A acid(6). The structures of compounds 1-6 were determined on the basis of physicochemical data analyses. Among them, compounds 1-3 exhibited a potent radical scavenging activity against 1,1-diphenyl-2-picrylhydrazyl(DPPH) with
values of 27.7, 23.4, and
Genetic and Antigenic Characterization of Swine H1N2 Influenza Viruses Isolated from Korean Pigs
Jo, Su-Kyoung ; Kim, Hyun-Soo ; Cho, Sung-Whan ; Seo, Sang-Heui ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 868~872
H1N2 influenza viruses are circulating in pigs worldwide and cause considerable economic losses to the pig industry. We genetically analyzed the genes of our isolates from Korean pigs, and compared the antigenicity of our isolates with swine H1N2 viruses isolated from pigs in the U.S.A. In addition, we serologically surveyed the infection rate of swine H1N2 viruses in pigs. We found that H1N2 isolates from Korean pigs are genetically more related to swine H1N2 viruses isolated from pigs in the U.S.A. than those in European countries. When antigenicity was compared, our isolates were weakly reacted to antibodies against swine H1N2 viruses isolated from pigs in the U.S.A. The serological surveillance using sera from pigs in Korea showed that about 46% was positive for H1N2 viruses. Our results suggest that swine H1N2 viruses are widespread in Korean pigs, and the development of a vaccine against H1N2 viruses may help to control their infection in pigs.
Microorganisms Against Plasmodiophora brassicae
Choi, Kwang-Hoon ; Yi, Yong-Sub ; Lee, Sun-Hee ; Kang, Kyung-Rae ; Lee, Eun-Jung ; Hong, Sung-Won ; Young, Jung-Mo ; Park, Young-Hee ; Choi, Gyung-Ja ; Kim, Bum-Joon ; Lim, Yoong-Ho ;
Journal of Microbiology and Biotechnology, volume 17, issue 5, 2007, Pages 873~877
In order to find microorganisms showing antifungal activities against Plasmodiophora brassicae, which causes club root, Korean salt-fermented fishery products were tested. Several fermented broths of microorgansims isolated from Ammodytes personatus fishery products showed high antifungal activities. The identification of microorganisms and their in vivo antifungal activities are reported herein.