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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 16, Issue 12 - Dec 2006
Volume 16, Issue 11 - Nov 2006
Volume 16, Issue 10 - Oct 2006
Volume 16, Issue 9 - Sep 2006
Volume 16, Issue 8 - Aug 2006
Volume 16, Issue 7 - Jul 2006
Volume 16, Issue 6 - Jun 2006
Volume 16, Issue 5 - May 2006
Volume 16, Issue 4 - Apr 2006
Volume 16, Issue 3 - Mar 2006
Volume 16, Issue 2 - Feb 2006
Volume 16, Issue 1 - Jan 2006
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Genetic Regulation of Corynebacterium glutamicum Metabolism
Wendisch Volker F. ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 999~1009
Physiological, biochemical and genetic studies of Corynebacterium glutamicum, a workhorse of white biotechnology used for amino acid production, led to a waste knowledge mainly about amino acid biosynthetic pathways and the central carbon metabolism of this bacterium. Spurred by the availability of the genome sequence and of genome-based experimental methods such as DNA microarray analysis, research on genetic regulation came into focus. Recent progress on mechanisms of genetic regulation of the carbon, nitrogen, sulfur and phosphorus metabolism in C. glutamicum will be discussed.
Effect of Viability and Integrity of Bifidobacterium on Suppression of Allergy in Mice
Kim Hye-Young ; Geun Eog-Ji ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1010~1016
The effects of the cell viability and integrity of Bifidobacterium on suppression of allergy were investigated. C3H/HeJ mice were sensitized on weeks 3, 4, 6, and 8 with ovalbumin and choleratoxin to induce an allergic reaction. Mice fed 0.2% of live, disrupted, or heat-killed Bifidobacterium bifidum BGN4 in the pellets of their diet for 8 weeks starting 2 weeks before initial sensitization differentially suppressed the allergy response in terms of levels of IgE and IgG1 in their sera, and symptoms on their tails. Viable Bifidobacterium was more effective than disrupted or heat-killed cells in suppressing the allergy. Growth inhibition, which occurred in the sham group at week 4, did not occur in the treated groups. These results show that Bifidobacterium has a suppressive effect on the allergic response of mice, and that the viability and integrity of the Bifidobacterium is required for effective suppression in our experimental model.
Generation and Segregation of Hantaviral RNA Genomic Diploid; Implications of Reassortant Generation Mechanism
Park Sun-Whan ; Chung Dong-Hoon ; Ahn Byung-Yoon ; Lee Pyung-Woo ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1017~1025
Hantaviruses possess three RNA segments of negative sense. Co-infection of closely related hantaviruses may result in generation of a progeny virus with genomic polyploidy, containing a partial or complete set of genome originated from more than one parental virus. To characterize the formation of viral genomic polyploidy, cultured Vero-E6 cells were co-infected with two closely related hantaviruses, Hantaan and Maaji, and the progeny viruses examined. The genotype of plaque-purified viruses was analyzed by a virus-specific RT-PCR. Seventy percent (67/96) of the progeny virus was categorized as Hantaan and 3.3% (2/96) was classified as Maaji, whereas 20% (21/96) was considered polyploidy as they contained both types of the S RNA segment. Most of the polyploidy progeny viruses were unstable and gave rise to either one of the parental viruses or a reassortant after several rounds of plaque purification. No recombination between the heterologous pair of S RNA was observed for those polyploid viruses during three consecutive plaque-to-plaque passages. These data suggest that the viral polyploidy formation constitutes a primary mechanism underlying the generation of a newly emerged hantavirus.
Production of a Fusion Protein Containing the Antigenic Domain 1 of Human Cytomegalovirus Glycoprotein B
Sousa Fani ; Ferreira Susana ; Queiroz Joao ; Domingues Fernanda ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1026~1031
The optimization of the production of a fusion protein containing the antigenic domain 1 (AD-1) is of a great importance, considering its use in diagnostic tests. The fusion protein is produced by the fermentation of a recombinant strain of Escherichia coli containing the plasmid Mbg58, which expresses the AD-1 (aa 484-650) of human cytomegalovirus glycoprotein B as a fusion protein together with aa 1-375 of
. An important characteristic of promoters (lac and derivatives) used in recombinant protein production in E. coli is their inducibility. Induction by IPTG is widely used for basic research; however, its use in large-scale production is undesirable because of its high cost and toxicity. In this work, studies using different inducers and carbon sources for the production of a fusion protein containing the AD-l were performed. The results showed that lactose could be used as an inducer in the fermentation process for the production of this protein, and that expression levels could exceed those achieved with IPTG. The use of lactose for protein expression in E. coli should be extremely useful for the inexpensive, large-scale production of heterologous proteins in E. coli. Addition of sucrose to the fermentation medium improved the yield of recombinant protein, whereas addition of fructose or trehalose decreased the yield.
Characterization of Styrene Catabolic Genes of Pseudomonas putida SN1 and Construction of a Recombinant Escherichia coli Containing Styrene Monooxygenase Gene for the Production of (S)-Styrene Oxide
Park Mi-So ; Bae Jong-Won ; Han Ju-Hee ; Lee Eun-Yeol ; Lee Sun-Gu ; Park Sung-Hoon ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1032~1040
Some Pseudomonas species can grow on styrene as a sole carbon and energy source. From the new isolate Pseudomonas putida SN1, the genes for styrene catabolism were cloned and sequenced. They were composed of four structural genes for styrene monooxygenase (styA and styB), styrene oxide isomerase (styC), and phenylacetaldehyde dehydrogenase (styD), along with two genes for the regulatory system (styS and styR). All the genes showed high DNA sequence (91% to 99%) and amino acid sequence (94% to 100%) similarities with the corresponding genes of the previously reported styrene-degrading Pseudomonas strains. A recombinant Escherichia coli to contain the styrene monooxygenase from the SN1 was constructed under the control of the T7 promoter for the production of enantiopure (S)-styrene oxide, which is an important chiral building block in organic synthesis. The recombinant E. coli could convert styrene into an enantiopure (S)-styrene oxide (ee >99%) when induced by IPTG The maximum activity was observed as 140 U/g cell, when induced with 1 mM IPTG at
Production of Selenium Peptide by Autolysis of Saccharomyces cerevisiae
Lee Jung-Ok ; Kim Young-Ok ; Shin Dong-Hoon ; Shin Jeong-Hyun ; Kim Eun-Ki ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1041~1046
Selenium-containing peptide (selenium peptide) was produced by autolysis of total proteins of Saccharomyces cerevisiae grown with inorganic selenium. Selenium peptide exhibited antioxidant activity as a glutathione peroxidase (GPx) mimic, and its activity was dependent on the hydrolysis methods. The GPx-like activity of the hydrolyzed selenium peptide increased 2.7-folds when digested by protease, but decreased by acid hydrolysis. During the autolysis of the yeast cell, the GPx-like activity and selenium content increased 4.3- and 2.3-folds, respectively, whereas the average molecular weight (MW) of selenium peptide decreased 70%. The GPx-like activity was dependent on the MW of selenium peptide and was the highest (220 U/mg protein) at 9,500 dalton. The maximum GPx-like activity (28,600 U/g cell) was obtained by 48 h of autolysis of the cells, which were precultured with 20 ppm of selenate. Selenium peptide showed little toxicity, compared with highly toxic inorganic selenium. These results show the potential of selenium peptide as a nontoxic antioxidant that can be produced by simple autolysis of yeast cells.
Purification and Characterization of Bile Salt Hydrolase from Lactobacillus plantarum CK 102
Ha Chul-Gyu ; Cho Jin-Kook ; Chai Young-Gyu ; Ha Young-Ae ; Shin Shang-Hun ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1047~1052
A bile salt hydrolase (BSH) was purified from Lactobacillus plantarum CK 102 and its enzymatic properties were characterized. This enzyme was successfully purified using ion-exchange chromatography with Q-Excellose and hydrophobic interaction chromatography with Butyl-Excellose. The purified enzyme showed a single protein band of 37 kDa by SDS-polyacrylamide gel electrophoresis, which was similar to the molecular weight of known BSHs. The amino acid sequence of GLGLPGDLSSMSR, determined by MALDI-TOF, was identical to that of BSH of L. plantarum WCFS1. Although this BSH hydrolyzed all of the six major human bile salts, glycine-conjugated bile acid was the best substrate, based on its specificity and
value. Among the various substrates, the purified enzyme maximally hydrolyzed glycocholate with apparent
values of 0.5 mM and 94 nmol/min/mg, respectively. The optimal pH of the enzyme ranged from 5.8 to 6.3. This enzyme was strongly inhibited by thiol enzyme inhibitors such as iodoacetate and periodic acid.
Film-Forming Properties of Proteinaceous Fibrous Material Produced from Soybean Fermented by Bacillus natto
Park Sang-Kyu ; Bae Dong-Ho ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1053~1059
The effectiveness of a proteinaceous fibrous material formed during commercial fermentation of soy protein (PFSP) and cysteine addition were evaluated in order to improve on the properties of soy protein-based films. Nine types of films were prepared at pH 7, 9, and 11, with heat treatments at
for 30 min, by casting 5% (w/w) PFSP aqueous solution, containing 2.25% (w/w) glycerol, on to polystyrene plates. The tensile strength (TS) of films ranged from 3.88 to 6.87 MPa. The highest puncture strength (PS) was observed with pH 7.0 films prepared from PFSP solution heated at
(P<0.05). Alkaline pH and temperature caused a decrease in both the TS and PS of the films. The thickness of films ranged from
. Water vapor permeabilities of the films decreased with increasing pH and temperature. To produce films from PFSP, pH value of 7.0 to 9.0 and heat treatment of
were needed. A soluble nature of PFSP films in water might be useful for preparation of hot water-soluble pouches. Cysteine addition could be necessary to produce films with increased TS and enhanced barrier properties. The combination treatment that provided the best combination of barrier and mechanical properties was the PFSP film prepared at pH 7.0 with addition of 1% cysteine. The films were good oxygen barriers.
Effect of Reservoirs on Microbiological Water Qualities in a Drinking Water Distribution System
Lee Dong-Geun ; Kim Sang-Jong ; Park Seong-Joo ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1060~1067
This study was undertaken to determine the effect of reservoirs on water quality and the distribution of pathogenic and indicator bacteria in a drinking water distribution system (total length 14km). Raw water, disinfected water, and water samples from the distribution system were subjected to physicochemical and microbiological analyses. Most factors encountered at each season included residual chloride, nitrate, turbidity, and phosphorus for heterotrophic bacterial distribution, and hardness, heterotrophic bacteria, sampling site, and DOC (dissolved organic carbon) for bacteria on selective media. No Salmonella or Shigella spp. were detected, but many colonies of opportunistic pathogens were found. Comparing tap water samples taken at similar distances from the water treatment plant, samples that had passed through a reservoir had a higher concentration of heterotrophic bacteria, and a higher rate of colony formation with 10 times as many bacteria on selective media. Based on the results with m-Endo agar, the water in reservoirs appeared safe; however, coliforms and opportunistic pathogenic bacteria such as Pseudomonas aeruginosa were identified on other selective media. This study illustrates that storage reservoirs in the drinking water distribution system have low microbiological water quality by opportunistic pathogens, and therefore, water quality must be controlled.
Plasmid- and Chromosome-Mediated Assimilation of Phenol and Cyanide in Pseudomonas sp. Strain PhCN
El-Deeb Bahig A. ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1068~1077
Pseudomonas sp. PhCN strain, which has the potential to utilize phenol and cyanide as a sole carbon and nitrogen source, was isolated. A comparison of the effect of cyanide on phenol degradation and vice versa by strain PhCN showed that the degradation time was significantly delayed by an increase in either phenol or cyanide concentration, and the greatest activities were obtained in basal medium containing a low concentration of cyanide and phenol. This strain contained two plasmids of approximately 120 kb (pPhCN-1) and 110 kb (pPhCN-2). Plasmid curing experiments produced a plasmid-free strain as well as strains containing either the 120- or the 110 kb plasmid. The strains were tested for their ability to utilize phenol and KCN. The results demonstrated that the ability to utilize phenol was encoded by the 120 kb plasmid, whereas the ability to utilize cyanide appeared to be encoded by the chromosome.
Inhibitory Effect of Capsaicin on Interleukin-8 Production by Helicobacter pylori-Infected MKN-45 Cells
Lee, Kwang-Hyoung ; Lee, Yong-Chan ; Kim, Tae-Il ; Noh, Sung-Hoon ; Kim, Ji-Yeon ; Paik, Hyun-Dong ; Kim, Chang-Han ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1078~1083
Capsaicin is the active ingredient in chili pepper and has an inhibitory effect on Helicobacter pylori growth and
activation. The present study examined the effect of capsaicin on interleukin (IL)-8 production by H. pylori ATCC 43504-infected MKN-45 cells, a gastric epithelial cell line. The viability of the MKN-45 cells treated with capsaicin at 0, 50, 100, 250, and
was 99, 98, 99, 99, and 85%, respectively. A capsaicin concentration as low as
significantly inhibited the IL-8 production induced by H. pylori ATCC 43504 infection (43.2% of control) during 24 h of incubation. However, low concentrations of capsaicin
did not significantly inhibit the IL-8 production by
or PMA-treated MKN-45 cells. Therefore, the overall inhibitory effect of capsaicin on H. pylori ATCC 43504 was the sum of H. pylori ATCC 43504 growth inhibition, host cell survival, and
signal cascade inhibition.
Lactic Acid Bacteria Increases Hypolipidemic Effect of Crocin Isolated from Fructus of Gardenia jasminoides
Lee In-Ah ; Min Sung-Won ; Kim Dong-Hyun ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1084~1089
The fructus of Gardenia jasminoides Ellis (GF) has been widely used as a natural colorant in Asian countries, and also as a Chinese traditional medicine for its homeostatic, antiphlogistic, analgesic, and antipyretic effects. In the present study, its main component, crocin, was fermented with lactic acid bacteria (LAB) and their antihyperlipidemic activity was measured. The GF extract, fermented GF (F-GF), crocin, and fermented crocin (F-crocin) significantly inhibited the increase of serum triglyceride (TG) level in corn oil feeding-induced triglyceridemic mice, as well as that of serum TG and total and LDL cholesterol levels in Triton WR-1339-induced hyperlipidemic mice. These agents also showed hypolipidemic activity in hyperlipidemic mice induced by high fat diet for 5 weeks. F-GF and F-crocin exhibited more potent hyperlipidemic effects than GF and crocin. The results suggest that the hypolipidemic effect of GF and crocin can be synergistically activated by LAB, and that F-GF and F-crocin may improve hyperlipidemia in clinic, compared with GF and crocin.
Biotransformation of Flavonoids with O-Methyltransferase from Bacillus cereus
Lee Yoon-Jung ; Kim Bong-Gyu ; Park Young-Hee ; Lim Yoong-Ho ; Hur Hor-Gil ; Ahn Joong-Hoon ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1090~1096
O-Methylation is a common modification reaction found in nature, and is mediated by an O-methyltransferase (OMT). OMTs have been mainly studied in plants, whereas only a few OMTs have been studied in microbes. When searching the Bacillus cereus genome, four putative small molecular OMTs were identified, among which BcOMT-1 was cloned and expressed in E. coli as a his-tag fusion protein. The whole cell expressing BcOMT-1 was used to methylate several flavonoids. Eriodictyol, luteolin, quercetin, and taxifolin, all of which contain 3' and 4' hydroxyl groups, served as methyl group acceptors for BcOMT-1, whereas naringenin, apigenin, 3,3'-dihydroxyflavone, and 3,4'-dihydroxyflavone did not function as substrates. Analysis of the reaction products using HPLC showed two different peaks, and NMR revealed that the methylation position was at the hydroxyl group of either carbon 3' or 4'. Therefore, this showed that BcOMT-1 used flavonoids containing ortho hydroxyl groups and transferred a methyl group to either of two hydroxyl groups.
Increase of Spacer Sequence Yields Higher Dimer
Yoo Mee-Hyeon ; Won Jae-Seon ; Lee Yong-Chan ; Choe Mu-Hyeon ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1097~1103
The divalent antibody-toxins are expected to have increased binding avidities to target cells because of the two cell-binding domains. However, previous studies showed that the refolding yield of divalent antibody-toxin is very low, and it is assumed that homodimer formation of antibody-toxin is strongly interfered by the repulsion between the two large toxin domains that come close to each other during dimer formation. In this study, B3 antibody was used as a model antibody, and its Fab domain was used to construct three different kinds of Fab divalent molecules,
. The monomer Fab-toxin molecules were made by fusing the Fab domain of monoclonal antibody B3 to PE38, a truncated mutant form of Pseudomonas exotoxin (PE), and a connecting sequence that contained spacer amino acid sequence (G4S)n (n=l, 2, 3) was inserted between Fab and PE38. The prepared divalent molecules were
, and they are derivatives of previously studied
, two Fab-S1, 2, 3-PE38 monomers were covalently linked by the disulfide bond bridge made from cysteine in the -SKPCIST- sequence. The insertion of spacer amino acids after the disulfide bridge resulted in a 12-18 fold higher yield of dimer formation than previously constructed
, 3-4-fold higher than
. These two molecules have less amino acid spacer sequence between the disulfide bridge and PE38 domain. The design of
in this study gave molecules with a higher refolding yield. The results of cytotoxicity assay showed a higher cytotoxic effect of these divalent molecules than that of the monovalent scFv-PE38 molecule.
Increased Refolding Yield of Disulfide Bond Bridged Fab-Toxin Homodimers by the Insertion of CH3 Domains
Song Jeong-Wha ; Won Jae-Seon ; Lee Yong-Chan ; Choe Mu-Hyeon ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1104~1110
Recombinant antibody-toxin is a bifunctional protein that binds and kills a target cell expressing a specific antigen on the surface of the cell, and its structure is chimeric, in which a toxin is fused to an antigen-binding domain such as scFv or Fab. Divalent antibody-toxin molecules showed higher cytotoxicities against cancer cell lines than monovalent molecules. However, the yields of the divalent molecules were very low. In this study, we introduced the CH2, CH3, or CH2-CH3 (=Fc) domain of antibody in the middle of the Fab-toxin between the hinge region of human IgG1 and the toxin domain to increase the yield. The covalently bonded dimer could be formed by three disulfide bridges from cysteine residues in the hinge region. The molecule with the CH3 domain showed about 3-fold higher dimerization yield than previously constructed Fab-toxin molecules, while maintaining the cytotoxic activity comparable to that of scFv-toxin. However, the introduction of CH2 or Fc domain to the same position showed little effect on the dimerization yield. We also observed that the introduction of the CH3 region made it possible to form noncovalently associated dimer molecules.
Analysis of Beauvericin and Unusual Enniatins Co-Produced by Fusarium oxysporum FB1501 (KFCC 11363P)
Song Hyuk-Hwan ; Ahn Joong-Hoon ; Lim Yoong-Ho ; Lee Chan ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1111~1119
Beauvericins and enniatins are cyclohexadepsipeptides exhibiting various biological activities on animal systems, including humans. Fusarium oxysporum FB1501 (KFCC 11363P) that produces four different cyclohexadepsipeptides was isolated from soil in Korea and the structures of the four cyclohexadepsipeptides elucidated by HPLC, MS, IR, and NMR analyses. The molecular weights for compounds 1,2,3, and 4 were determined to be 654.5, 784.5, 668.6, and 682.5, respectively, on the basis of ESI-MS measurements. The IR spectra for all the compounds exhibited absorptions for ester
bonds that were very similar to those for beauvericin and enniatins with ester and amide absorptions. The results of the NMR analysis
revealed that compounds 1,3, and 4 consisted of
(Hiv), and 2-hydroxy-3-methylpentanoic acid (Hmp) residues (compound 1: three N-MeVal residues, two Hiv residues, and one Hmp residue; compound 3: three N-MeVal residues, one Hiv, and two Hmp residues; compound 4: three N-MeVal residues and three Hmp residues). Therefore, the compounds were identified as enniatin H (compound 1), enniatin I (compound 3), and enniatin MK1688 (compound 4). Compound 2 was analyzed as beauvericin according to 1D and 2D NMR analyses. This study is the first report related to the co-production of beauvericin with other unusual enniatins, such as enniatin H, enniatin I, and enniatin MK1688, by Fusarium oxysporum.
Stimulation of Cephalosporin C Production by Acremonium chrysogenum M35 with Fatty Acids
Kim Jong-Chae ; Kang Seong-Woo ; Lim Jung-Soo ; Song Yoon-Seok ; Kim Seung-Wook ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1120~1124
Supplementation with rice oil and its major components (oleic acid and linoleic acid) was found to have a significant influence on cephalosporin C (CPC) production and cell growth by A. chrysogenum M35 in shake flask cultures. Five percent (v/v) rice oil had the most robust effect and 5% (v/v) oleic acid was the second most efficient on cell growth, whereas 3% (v/v) linoleic acid was found to be optimal for CPC production. Rice oil, oleic acid, and linoleic acid also significantly improved the rates of glucose consumption. When glucose was almost consumed, CPC production was initiated and, on the addition of rice oil, lipase activity increased steadily to 1.56 U/ml for 4 days. These results suggest that rice oil and fatty acids are used as carbon source to produce CPC by A. chrysogenum M35. Moreover, a mixture, composed of 40% (v/v) oleic acid and 60% (v/v) linoleic acid, had the strongest stimulatory effect on CPC production, due to a synergistic effect of the two fatty acids. Consequently, the maximum CPC titer (7.44 g/l) was improved about 4.5-fold.
Induction of Kanamycin Resistance Gene of Plasmid pUCD615 by Benzoic Acid and Phenols
Mitchell Robert J. ; Hong Han-Na ; Gu Man-Bock ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1125~1131
A kan'::luxCDABE fusion strain that was both highly bioluminescent and responsive to benzoic acid was constructed by transforming E. coli strain W3110 with the plasmid pUCDK, which was constructed by digesting and removing the 7-kb KpnI fragment from the promoterless luxCDABE plasmid pUCD615. Experiments using buffered media showed that this induction was dependent on the pH of the media, which influences the degree of benzoic acid protonation, and the expression levels seen are likely due to acidification of the cytoplasm by uncoupling of benzoic acid. Consequently, the sensitivity of this strain for benzoic acid was increased by nearly 20-fold when the pH was shifted from 8.0 to 6.5. Benzoic acid derivatives and several phenolics also resulted in significantly increased bioluminescent signals. Although these compounds are known to damage membranes and induce the heat-shock response within E. coli, bacterial strains harboring mutations in the fadR and rpoH genes, which are responsible for fatty acid biosynthesis during membrane stress and induction of the heat-shock response, respectively, showed that these mutations had no effect on the responses observed.
Purification and Characterization of Novel Bifunctional Xylanase, XynIII, Isolated from Aspergillus niger A-25
Chen Hong-Ge ; Yan Xin ; Liu Xin-Yu ; Wang Ming-Dao ; Huang Hui-Min ; Jia Xin-Cheng ; Wang Jin-An ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1132~1138
Three types of xylanases (EC 184.108.40.206) were detected in the strain Aspergillus niger A-25, one of which, designated as XynIII, also displayed
(EC 220.127.116.11) activity, as determined by a zymogram analysis. XynIII was purified by ultrafiltration and ion-exchange chromatography methods. Its apparent molecular weight was about 27.9 kDa, as estimated by SDS-PAGE. The purified XynIII could hydrolyze birchwood xylan, oat spelt xylan, lichenin, and barley
, but not CMC, avicel cellulose, or soluble starch under the assay conditions in this study. The xylanase and
activities of XynIII both had a similar optimal pH and pH stability, as well as a similar optimal temperature and temperature stability. Moreover, the effects of metal ions on the two enzymatic activities were also similar. The overall hydrolytic rates of XynIII in different mixtures of xylan and lichenin coincided with those calculated using the Michaelis-Menten model when assuming the two substrates were competing for the same active site in the enzyme. Accordingly, the results indicated that XynIII is a novel bifunctional enzyme and its xylanase and
activities are catalyzed by the same active center.
Accurate Metabolic Flux Analysis through Data Reconciliation of Isotope Balance-Based Data
Kim Tae-Yong ; Lee Sang-Yup ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1139~1143
Various techniques and strategies have been developed for the identification of intracellular metabolic conditions, and among them, isotope balance-based flux analysis with gas chromatography/mass spectrometry (GC/ MS) has recently become popular. Even though isotope balance-based flux analysis allows a more accurate estimation of intracellular fluxes, its application has been restricted to relatively small metabolic systems because of the limited number of measurable metabolites. In this paper, a strategy for incorporating isotope balance-based flux data obtained for a small network into metabolic flux analysis was examined as a feasible alternative allowing more accurate quantification of intracellular flux distribution in a large metabolic system. To impose GC/MS based data into a large metabolic network and obtain optimum flux distribution profile, data reconciliation procedure was applied. As a result, metabolic flux values of 308 intracellular reactions could be estimated from 29 GC/ MS based fluxes with higher accuracy.
Characterization of a Noncanonical Purine dNTP Pyrophosphatase from Archaeoglobus fulgidus
Im Eun-Kyoung ; Hong Chang-Hyung ; Back Jung-Ho ; Han Ye-Sun ; Chung Ji-Hyung ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1144~1148
DNA can oxidatively be deaminated by ROS, which converts DNA base amino groups to keto groups and can trigger abnormal mutations, resulting in mutagenesis in organisms. In this study, a noncanonical purine dNTP pyrophosphatase (AfPPase) from a hyperthermophilic archaeon Archaeoglobus fulgidus, which hydrolyzes aberrant nucleoside triphosphates, was overexpressed in E. coli, purified, and characterized. The purified AfPPase showed remarkably high activity for XTP and dITP, suggesting that the 6-keto group of these nucleotides is critical for the reactivity. Under optimal reaction conditions, the reaction rate for these substrates was about 120 times that with dGTP. Therefore, AfPPase may play a significant role in DNA repair by hydrolysis of noncanonical nucleotides before they are misincorporated into DNA.
In Vitro Selection of Cancer-Specific RNA Aptamers
Lee Young-Ju ; Lee Seong-Wook ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1149~1153
In this study, nuclease-resistant RNA aptamers that are specific for Jurkat T leukemia cells were selected by a subtractive systemic evolution of ligands by exponential enrichment (SELEX) method. A randomized nuclease-resistant RNA library was incubated with normal peripheral blood mononuclear cells (PBMC) in each round to preclude RNAs that recognize the common cellular components on the surface of normal and cancer cells. The precluded RNAs were used for the selection of Jurkat T cell-specific aptamers, and the specific RNAs were then gradually enriched from start to the following selections. After 16 rounds of the subtractive SELEX, the selected aptamers were found to preferentially bind to Jurkat T cells, but not to the normal PBMC, evidenced by fluorescence-activated cell sorting analysis. Thus, the subtractive SELEX can be used to identify ligands to cancer-specific biological markers without prior knowledge of the nature of markers. The aptamers could be applied to specific cell sorting, tumor therapy, and diagnosis, and moreover, to find cancer cell-specific markers.
S-Adenosyl-L-Methionine Analogues to Enhance the Production of Actinorhodin
Chong You-Hoon ; Young Jung-Mo ; Kim Jin-Young ; Lee Yu-Kyung ; Park Kwang-Su ; Cho Jun-Ho ; Kwon Hyung-Jin ; Suh Joo-WOn ; Lim Yoong-Ho ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1154~1157
It is known that overexpression of S-adenosyl-L-methionine (SAM) synthetase or exogenous addition of SAM enhances the production of actinorhodin, one of pigmented antibiotics found from Streptomyces coelicolor. In order to discover a novel compound as a signal molecule to produce actinorhodin instead of SAM, several compounds were synthesized based on the relationships between structures of the SAM analogues and their actinorhodin productivities. Of these, a few compounds showed better productivities of actinorhodin than SAM.
Effects of Isocitrate Lyase Inhibitors on Spore Germination and Appressorium Development in Magnaporthe grisea
Kim Seung-Young ; Park Jin-Soo ; Oh Ki-Bong ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1158~1162
The glyoxylate cycle can conserve carbons and adequately supply tricarboxylic acid (TCA) cycle intermediates for biosynthesis when microorganisms grow on
carbon sources. It has been reported that isocitrate lyase (ICL1), a key enzyme of the glyoxylate cycle, is highly induced when Magnaporthe grisea, the causal agent of rice blast, infects its host. Therefore, the glyoxylate cycle is considered as a new target for antifungal agents. A 1.6-kb DNA fragment encoding the ICL1 from M. grisea KJ201 was amplified by PCR, cloned into a vector providing His-tag at the N-terminus, expressed in Escherichia coli, and purified using Ni-NTA affinity chromatography. The molecular mass of the purified ICL1 was approximately 60 kDa, as determined by SDS-PAGE. The ICL1 inhibitory effects of TCA cycle intermediates and their analogs were investigated. Among them, 3-nitropropionate was found to be the strongest inhibitor with an
. 3-Nitropropionate inhibited the appressorium development in M. grisea at the
level, whereas conidia germination remained unaffected. This compound also inhibited the mycelial growth of the fungus on minimal medium containing acetate as a
carbon source. These results suggest that ICL1 plays a crucial role in appressorium formation of M. grisea and is a new target for the control of phytopathogenic fungal infection.
Cloning, Sequencing and Characterization of Acyltransferase Gene Involved in Exopolysaccharide Biosynthesis of Zoogloea ramigera 115SLR
Lee Sam-Pin ; Troyano Esperanza ; Lee Jin-Ho ; Kim Hyun-Soo ; Sinskey Anthony John ;
Journal of Microbiology and Biotechnology, volume 16, issue 7, 2006, Pages 1163~1168
The recombinant plasmid pLEX2FP complements the mutation in Zoogloea ramigera 115MM1, and the complemented mutant produces an exopolysaccharide that shows higher affinity for the calcofluor dye than the exopolysaccharide from Z. ramigera 115SLR, resulting in higher fluorescence intensity under UV light. A compositional and structural analysis of the exopolysaccharide from Z. ramigera 115MM1 showed that the different fluorescent properties were due to a lower content of acetyl groups when compared with Z. ramigera 115SLR exopolysaccharide. These results were in agreement with a sequence analysis of the gene carried in the plasmid pLEX2FP, which appeared to encode an O-acyltransferase highly homologous to the 3-O-acyltransferase of Streptomyces mycarofaciens. The gene encoding the acyltransferase from Z. ramigera 115SLR was expressed as a GST-fusion protein with 70,000 daltons in E. coli.