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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
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Journal of Microbiology and Biotechnology
Journal Basic Information
Journal DOI :
The Korean Society for Applied Microbiology and Biotechnology
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Volume & Issues
Volume 7, Issue 6 - Dec 1997
Volume 7, Issue 5 - Oct 1997
Volume 7, Issue 4 - Aug 1997
Volume 7, Issue 3 - Jun 1997
Volume 7, Issue 2 - Apr 1997
Volume 7, Issue 1 - Feb 1997
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Gene Amplification of aceA and aceB in Lysine-producing Corynebacterium glutamicum ssp. lactofermentum ATCC21799
Kim, Hyung-Joon ; Kim, Youn-Hee ; Lee, Heung-Shick ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 287~292
The role of glyoxylate bypass in lysine production by Corynebacterium glutamicum ssp. lactofermentum ATCC21799 was analyzed by using cloned aceA and aceB genes which encode enzymes catalyzing the bypass. Introduction of a plasmid carrying aceA and aceB to the strain increased enzyme activities of the bypass to approximately 5 fold on acetate minimal medium. The strain with amplified glyoxylate bypass excreted 25% more lysine to the growth medium than the parental strain, apparently due to the increased availability of intracellular oxaloacetate. The final cell yield was lower in the strain with amplified glyoxylate bypass. These changes were specific to the lysine-producing C. glutamicum ssp. lactofermentum ATCC21799, since the lysine-nonproducing wild type Corynebacterium glutamicum strain grew faster and achieved higher cell yield when the glyoxylate bypass was amplified. These findings suggest that the lysine producing C. glutamicum ssp. lactofermentum ATCC21799 has the ability to efficiently channel oxaloacetate, the TCA cycle intermediate, to the lysine biosynthesis pathway whereas lysine-nonproducing strains do not. Our results show that amplification of the glyoxylate bypass efficiently increases the intracellular oxaloacetate in lysine producing Corynebacterium species and thus results in increased lysine production.
Cloning of the Entire Gene Encoding the 140-kDa
-Amylase of Lactobacillus amylovorus and Expression in Escherichia coli and Lactococcus lactis
Jeong, Jong-Jin ; Kim, Tea-Youn ; Kim, Jeong-Hwan ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 293~298
A 4.6-kb HindIII fragment encompassing the complete 140-kDa
-amylase gene of Lactobacillus amylovorus B 4540 was cloned into pBR322 by the shot gun method. Southern blotting and restriction mapping for the insert were performed. The recombinant 9.0-kb plasmid, pFML1, conferred
-amylase activity to E. coli and Lactococcus lactis hosts when introduced by electroporation. SDS-PAGE and zymography confirmed the production of 140-kDa
-amylase and its proteolytic degradation products with enzyme activity in transformants. Total
-amylase activity of E. coli
cells harboring pFML1 was 1.8 units and most activity was detected from cell pellets. Total enzyme activity of L. lactis subsp. lactis MG1363 transformant was five to ten-fold lower than that of E. coli cell but more than half of the activity was detected in the culture supernatant.
Periplasmic Expression of a Recombinant Antibody (MabB9) in Escherichia coli
Chang, Hae-Choon ; Kwak, Ju-Won ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 299~304
Expression in the periplasm of Escherichia coli of cloned heavy and light chain cDNAs for Fab fragment of a murine monoclonal antibody MabB9 (
, K), specific for human plasma apolipoprotein B-100 of LDL, was studied. For the purpose, a vector for two-cistronic expression of the heavy chain cDNA, at the 5' terminus, and light chain cDNA, at the 3' terminus, was constructed using the signal sequences, pelB (for heavy chain) and ompA (for light chain) in a pET vector system. The constructed vector was transformed into E. coli BL21(DE3). The expressed heavy chain (25 kDa) and light chain (23 kDa) of the antibody molecule were detected in total cell extracts as well as in the periplasmic extracts of E. coli.
Purification and Characterization of Carboxymethyl Cellulase from Bacillus stearothermophilus No. 236
Kim, Sohng-Hwan ; Cho, Ssang-Goo ; Choi, Yong-Jin ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 305~309
Bacillus stearothermophilus No. 236, an effective xylanolytic bacterium, produced an extracellular carboxymethyl cellulase when the strain was grown on xylan. The carboxymethyl cellulase was purified to homogeneity as judged by SDS-PAGE and zymogram, The carboxymethyl cellulase had a pI of 4.0, and a molecular mass of 95 kDa. The highest level of enzyme activity was observed at pH 6.5 and
values of the enzyme to carboxymethyl cellulose were 20.8 mg/ml and
/min/mg protein, respectively, The enzyme was found to act also on filter paper and xylan as well as carboxymethyl cellulose. Therefore, it is expected that this xylanolytic strain isolated from soil could be efficiently used for xylan biodegradation.
Synthesis of Glucosyl-sugar Alcohols Using Glycosyltransferases and Structural Identification of Glucosyl-maltitol
Kim, Tae-Kwon ; Park, Dong-Chan ; Lee, Yong-Hyun ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 310~317
Enzymatic synthesis of glucosyl-sugar alcohols using various transglycosylating enzymes, such as cyclodextrin glucanotransferase (CGTase),
-glucosidase, and pullulanase was investigated using various sugar alcohols, such as sorbitol, xylitol, inositol, maltitol, and lactitol as glucosyl acceptors. CGTase showed the highest transglycosylating activity to sugar alcohols compared to other transglycosylating enzymes, and inositol and maltitol were the most suitable glucosyl acceptors. Soluble starch, extruded starch, cyclodextrins, and maltooligosaccharides were also identified to be adequate glucosyl donors for transglycosylation reaction of CGTase to sugar alcohols. The synthesis of glucosyl-maltitol in the reaction system using extruded starch as the glucosyl donor and maltitol as the glucosyl acceptor showed the best results showing the highest transglycosylation yield. The transglycosylation products were purified by activated carbon column chromatography with ethanol gradient elution. Chemical structures of above transglucosylated products were analyzed by nuclear magnetic resonance spectroscopy, and two products were identified to be maltotritol and maltotetraitol, in which one or two glucose molecules attached to the parent maltitol molecule by a
-l,4-glucosidic bond, respectively.
Optimal Resolution of L-Carnitine from Racemic DL-Carnitine by Enterobacter sp. Assimilating D-Carnitine
Hwang, Ki-Chul ; Bang, Won-Gi ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 318~322
In order to isolate a microorganism having preferential degradation of D-carnitine from DL-carnitine, a bacterium assimilating D-carnitine as a sole carbon and energy source was isolated from soil by enrichment culture and partially identified as Enterobacter sp. Also, a mutant having lessened L-carnitine decomposition rates was selected with nitrosoguanidine mutagenesis, which led to decrease the specific activities of carnitine dehydrogenase (7.6-fold) and
-hydroxybutyrate dehydrogenase (9.5-fold) as compared to the wild strain. Meanwhile, optimal culture conditions for optical resolution of DL-carnitine were investigated. Under optimal conditions, 3.53 g/l L-carnitine was obtained from 20 g/l DL-carnitine, which corresponded to 35.3% L-carnitine yield and 97.9% optical purity.
Fed-batch Fermentations of Recombinant Escherichia coli to Produce Bacillus macerans CGTase
Park, Yong-Cheol ; Kim, Chang-Sup ; Kim, Chung-Im ; Choi, Kyu-Hwan ; Seo, Jin-Ho ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 323~328
The recombinant Escherichia coli BL21(DE3)pLysE : pTCGT1 was grown to overproduce Bacillus macerans cyclodextrin glucanotransferase (CGTase) able to synthesize
-cyclodextrin (CD) with a selectivity of 67%. A number of batch fermentations were performed to test the possibility of using lactose as an inducer of the E. coli T7 promoter system. A mixture of isopropyl
-D-thiogalactoside (IPTG) and lactose (1 : 1) gave a maximum CGTase activity of 2.4 U/ml, which was higher than the value obtained with induction by IPTG alone. Fed-batch fermentations involving a glucose-controlled growth period followed by a gene-expression phase with mixtures of IPTG and lactose were employed to achieve high cell density and thereby increase total CGTase activity. Optimized fed-batch fermentation using the modified inducer (IPTG : lactose=1 : 3) and 100 g/l yeast extract solution in the gene-expression phase resulted in a maximum CGTase activity of 62.9 U/ml and a final cell mass of 53.5 g/l, corresponding to a 31-fold increase in CGTase activity and a 29-fold increase in cell mass compared with the control batch fermentation.
Enantioselective N-Acetylation of 3-Amino-3-phenylpropionic Acid by Cell-free Extracts of Streptomyces neyagawaensis
Chung, Myung-Chul ; Lee, Ho-Jae ; Lee, Choong-Hwan ; Chun, Hyo-Kon ; Kho, Yung-Hee ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 329~332
Cell-free extracts of Streptomyces neyagawaensis SL-387 grown on a chemically defined medium supplemented with DL-3-amino-3-phenylpropionic acid (APP) produced N-acetyl-APP (Ac-APP) in the presence of APP and acetyl coenzyme A. The APP obtained by acid hydrolysis of the Ac-APP was D-configuration:
, optical purity 92% enantiomeric excesses (ee). These results suggest that an N-acetyltransferase exists in the cell-free extract as a novel enzyme with specificity for D-APP.
Selection and Identification of a Strain KT-10 Producing the Cathepsin B Inhibitor
Han, Kil-Hwan ; Do, Jae-Ho ; Kim, Sang-Dal ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 333~340
An actinomycetes, KT-10 isolated from ginseng field in Kyongpook, Korea was selected based on its ability to produce a lysosomal cathepsin B inhibitor. The inhibitor purified from the culture supernatant of the isolate KT-10 showed strong inhibitory effects against cathepsin B as well as against papain when the activities were measured using synthetic substrate,
-N-benzyloxycarbonyl-L-Iysine p-nitrophenyl ester (CLN) or
-N-benzoyl-D,L-arginine 2-naphthylamide (BANA). The isolate KT-10 was identified as a species of Streptomyces based on its morphological characteristics and chemotaxonomic data. The TAXON program of Ward was used to identify Streptomyces sp. KT-10 as a strain of Streptomyces luteogriseus belong to cluster 18 of the genus Streptomyces with a Willcox probability 0.999388. The cathepsin B inhibitor was presumed to a novel material composed of a polyhydroxylamine.
Sustained Production of Amino Acids by Immobilized Analogue- resistant Mutants of a Cyanobacterium Anacystis nidulans BD-1
Bagchi, Suvendra Nath ; Rao, Nandula Seshgiri ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 341~344
Batch cultures of Anacystis nidulans BD-1 resistant to azaleucine and fluorotyrosine produced and liberated a wide range of amino acids, notably glutamic acid, alanine, phenylalanine, leucine, isoleucine, cysteine and methionine. Sustained liberation for prolonged periods was achieved after immobilization on calcium alginate and the net concentration in the medium was 0.18-0.2 g
. While acetohydroxy acid synthase in azaleucine-resistant mutant lost leucine- and isoleucine-sensitivity, fluorotyrosine-resistant strain turned phenylalanine activating. The activities of nitrate assimilating enzymes were also higher in the mutants and were relaxed from ammonium-repression. The metabolic adjustments involved in amino acid overproduction are discussed.
Optimization of Catechol Production Using Immobilized Resting Cells of Pseudomonas putida in Aqueous/organic Two-phase System
Chae, Hee-Jeong ; Yoo, Young-Je ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 345~351
An aqueous/organic two-phase reaction system was applied to the production of catechol using immobilized resting cells of Pseudomonas putida CY 400. Water/ethyl ether system was used because of high partition coefficient of catechol and thus to reduce the product inhibition and degradation. Among the tested immobilization carriers, polyacrylamide gel gave the highest catechol productivity. The immobilization seemed to protect the cells against solvent toxicity. From the simulation of reaction conditions based on two-phase models, it was found that there was an optimum acetate concentration at fixed benzoate and cell concentrations for the catechol productivity. A lower phase volume ratio (lower fraction of organic phase) gave a higher productivity. However, the substrate conversion was low at low phase volume ratio.
Measurement of Iron-dependence of pupA Promoter Activity by a pup-lux Bioreporter
Khang, Yong-Ho ; Yang, Zamin-K. ; Burlage, Robert-S. ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 352~355
The promoter region of the pupA gene of Pseudomonas putida WCS358 was fused with the structural genes for bioluminescence (luxCDABE) from Vibrio fischeri, and the resulting fusion plasmid harbored by the WCS358 host. The pup-lux fusion gene was then used for quantitative analysis of the iron-dependence of pupA promoter activity. Factors affecting bioluminescence produced by the pup-lux bioreporter were found to be cell activity, iron-chelator concentrations, Fe(III) concentrations, and nutrient components. Light production rates of the pup-lux bioreporter were inversely dependent upon iron molecules when
concentrations were between
in nutrient-poor minimal media, and between 0.1 and 10 mM in nutrient-rich complex media.
Activities of Oxidative Enzymes Related with Oxygen Tolerance in Bifidobacterium sp.
Shin, Soon-Young ; Park, Jong-Hyun ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 356~359
To study the relationship between oxygen tolerance and enzyme activity in the oxygen metabolism of bifidobacteria, the activities of catalase, superoxide dismutase (SOD), NADH oxidase and NADH peroxidase from six typical bifidobacteria and other bacteria were assayed by spectrophotometry. Catalase activity was hardly detected in any of the bifidobacteria tested. SOD activity was detected in every species including the Clostridium species. In particular SOD activity was notably high in the aerosensitive Bifidobacterium adolescentis. This fact indicates that SOD activity is not a critical factor to ensure aerotolerance. Aerosensitive B. adolescentis showed very low NADH oxidative enzyme activity whereas other aerotolerant bifidobacteria exhibited considerable activity for the enzymes. It seems that detoxification of
by NADH oxidative enzymes might be an important factor in improving for aerotolerant bifidobacteria survival rates in an oxygen environment.
Isolation and Characterization of
-Hydroxybutyrate Dehydrogenase- deficient Mutant of Rhodobacter sphaeroides 2.4.1
Kho, Dohng-Hyo ; Lee, Jeong-Kug ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 360~362
A transposon Tn5 mutant of Rhodobacter sphaeroides 2.4.1 was isolated for its impaired ability of growth on minimal medium containing
-hydroxybutyric acid as a sole carbon source. The mutant, R. sphaeroides S7 showed approximately 6-fold decrease in
-hydroxybutyrate dehydrogenase activity compared with that of wild type. In R. sphaeroides S7 the Tn5 was located in DNA region corresponding to a 4.2-kb EcoRI DNA fragment of R. sphaeroides 2.4.1 chromosome.
Characterization of Doxorubicin-nonproducing Mutant, Nu3 of Streptomyces peucetius ATCC27952
Kyu, Hwang-Cheol ; Lee, Hong-Sub ; Hong, Young-Soo ; Paek, Nam-Soo ; Kim, Tae-Han ; Lee, Jung-Joon ;
Journal of Microbiology and Biotechnology, volume 7, issue 5, 1997, Pages 363~366
A doxorubicin-nonproducing mutant, Nu23 was selected from the mutagenesis of Streptomyces peucetius ATCC27952. Chemical and molecular biological analysis suggested that the mutant was blocked at the step between polyketide synthase and aklaviketon reductase in the biosynthesis of doxorubicin. Furthermore, the bioconversion experiment with the mutant revealed that 13-dihydrodaunorubicin is likely to be a biosynthetic intermediate.