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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 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
Selecting the target year
Immunosensor for Detection of Escherichia coli O157:H7 Using Imaging Ellipsometry
Bae Young-Min ; Park Kwang-Won ; Oh Byung-Keun ; Choi Jeong-Woo ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1169~1173
Imaging ellipsometry (IE) for detection of binding of Escherichia coli O157:H7 (E. coli O157:H7) to an immunosensor is reported. A protein G layer, chemically bound to a self-assembled layer of 11-mercaptoundecanoic acid (11-MUA), was adopted for immobilization of monoclonal antibody against E. coli O157:H7 (Mab). The immobilization of antibody was investigated using surface plasmon resonance. To fabricate antibody spots on a gold surface, protein G solution was spotted onto the gold surface modified with an 11-MUA layer, followed by immobilizing Mab on the protein G spot. Ellipsometric images of the protein G spot, the Mab spot, and Mab spots with binding of E. coli O157:H7 in various concentrations were acquired using the IE system. The change of mean optical intensity of the Mab spots in the ellipsometric images indicated that the lowest detection limit was
CFU/ml for E. coli O157:H7. Thus, IE can be applied to an immunosensor for detection of E. coli O157:H7 as a detection method with the advantages of allowing label-free detection, high sensitivity, and operational simplicity.
Deregulation of Aspartokinase by Single Nucleotide Exchange Leads to Global Flux Rearrangement in the Central Metabolism of Corynebacterium glutamicum
Kim Hyung-Min ; Heinzle Elmar ; Wittmann Christoph ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1174~1179
The wild-type Corynebacterium glutamicum ATIC 13032 and Corynebacterium glutamicum ATTC 13032 lysC S301Y, exhibiting a deregulated aspartokinase, were compared concerning growth, lysine production, and intracellular carbon fluxes. Both strains differ by only one single nucleotide over the whole genome. In comparison to the wild-type, the mutant showed significant production of lysine with a molar yield of 0.087 mol (mol glucose
) whereas the biomass yield was reduced. The deregulation of aspartokinase further led to a global rearrangement of carbon flux throughout the whole central metabolism. This involved an increased flux through the pentose phosphate pathway (PPP) and an increased flux through anaplerosis. Because of this, the mutant revealed an enhanced supply of NADPH and oxaloacetate required for lysine biosynthesis. Additionally, the lumped flux through phosphoenolpyruvate carboxykinase and malic enzyme, withdrawing oxaloacetate back to the glycolysis and therefore detrimental for lysine production, was increased. The reason for this might be a contribution of malic enzyme to NADPH supply in the mutant in the mutant. The observed complex changes are remarkable, because they are due to the minimum genetic modification possible, the exchange of only one single nucleotide.
Identification and Cloning of a Fraction 1 Protein of Yersinia pestis that Produces Protective Immune Responses
Kim Jong-Hyun ; Cho Seung-Hak ; Jang Hyun-Chul ; Lee Hee-Cheul ; Kim Young-Il ; Kang Yeon-Ho ; Lee Bok-Kwon ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1180~1184
The capsule that surrounds Yersinia pestis cells is composed of a protein-polysacchride complex; the purified protein component is fraction I (F1) antigen. We report the cloning of the cafl gene and its expression in Escherichia coli using the vector pETl02/D-TOPO and the F1-specific monoclonal antibody. The recombinant F1 (rF1) antigen had a molecular size of 17.5 kDa, which was identical to that of the F1 antigen produced by Y. pestis. Recombinant F1 protein was found to react to polyclonal antiserum to Y. pestis Fl. Recombinant F1 was purified by ProBond purification system and induced a protective immune response in BALB/c mice challenged with up to 10
virulent Y. pestis. Purified rF1 protein was used in an ELISA to evaluate the ability of a method to detect antibodies to Y. pestis in animal sera. These results strongly indicated that the rF1 protein is a suitable species-specific immunodiagnostic antigen and vaccine candidate.
Cloning and Overexpression of Gene Encoding the Pullulanase from Bacillus naganoensis in Pichia pastoris
Xu Bo ; Yang Yun-Juan ; Huang Zun-Xi ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1185~1191
The expression of a pullulanase gene in Pichia pastoris was investigated. The gene encoding pullulanase was cloned by PCR using the chromosomal DNA of Bacillus naganoensis as the template. The expression vector pPIC9K-Pu was constructed by inserting the pullulanase gene into plasmid pPIC9K and then transformed into Pichia pastoris SMD 1168 by electroporation. Activity determination, SDS-PAGE, and PCR amplification indicated that the gene of the pullulanase from B. naganoensis had successfully been expressed in SMD 1168 and the molecular size of the expressed recombinant product was about 119.9 kDa. This is the first report on the successful expression of the pullulanase from B. naganoensis in P. pastoris. The transformant secreted recombinant pullulanase with the activity of 350.8 IU/ml in shake-flask culture. The properties of the recombinant pullulanase were characterized.
Characterization of an Apple Polygalacturonase-Inhibiting Protein (PGIP) That Specifically Inhibits an Endopolygalacturonase (PG) Purified from Apple Fruits Infected with Botryosphaeria dothidea
Lee Dong-Hoon ; Bae Han-Hong ; Kang In-Kyu ; Byun Jae-Kyun ; Kang Sang-Gu ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1192~1200
An apple polygalacturonase-inhibiting protein (PGIP), which specifically inhibits endopolygalacturonase (PG, EC 188.8.131.52) from Botryosphaeria dothidea, was purified from Botryosphaeria dothidea-infected apple (Malus domestica cv. Fuji) fruits. The purified apple PGIP had a molecular mass of 40 kDa. The N-terminal amino acid sequence of the purified protein showed high homologies to those of PGIP from pear (100%), tomato (70%), and bean (65%). We also purified polygalacturonase (PG) from B. dothidea. The PG hydrolyzes pectic components of plant cell walls. When the extracted apple pectic cell wall material was treated with purified apple PGIP and B. dothidea PG, the amount of uronic acid released was lower than that treated with B. dothidea PG alone. This result demonstrates that PGIP functions specifically by inhibiting cell wall maceration of B. dothidea PG Furthermore, we characterized the de novo function of the PGIP against PG on the solubilization and depolymerization of polyuronides from cell wall of apple fruits inoculated with B. dothidea. This result demonstrated that the PGIP of plants exhibits one of the direct defense mechanisms against pathogen attack by inhibiting PGs that are released from pathogens for hydrolysis of cell wall components of plants.
Cholesterol Lowering Effect of Lactobacillus plantarum Isolated from Human Feces
Ha Chul-Gyu ; Cho Jin-Kook ; Lee Chi-Ho ; Chai Young-Gyu ; Ha Young-Ae ; Shin Shang-Hun ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1201~1209
The purpose of this study was to isolate probiotic lactic acid bacteria (LAB) that produce bile salt hydrolase (BSH), and to evaluate its effects on serum cholesterol level. One-hundred-twenty bacterial colonies were initially isolated from human feces, and five strains were selected after screening based on their resistance to acids, tolerance against bile salts, and inhibitory activity on Escherichia coli. The Lactobacillus plantarum strain with the highest level of BSH activity was identified using 16S rRNA sequences, and was named L. plantarum CK 102. L. plantarum CK 102 at a level of 1.36
cfu/ml survived in pH 2 buffer for 6 h and exhibited excellent tolerance for bile salt. Coculturing the strain with E. coli in MRS broth resulted in strong inhibition against growth of E. coli at 18 h. Furthermore, the potential effect of CK 102 on serum cholesterol level was evaluated in rats. Thirty-two rats [Sprague-Dawley (SD) male, 129
l g, 5 weeks old] were divided into four groups of eight each. For six weeks, Group 1 was fed a normal diet (negative control); Group 2 was fed a cholesterol-enriched diet (positive control); Group 3 was fed a cholesterol-enriched diet plus L. plantarum CK 102 at 1.0
cfu/ml; and Group 4 was fed a cholesterol-enriched diet plus L. plantarum CK 102 at 5.0
cfu/ml. Blood samples were collected, serum lipids were analyzed, and weights of the organs were measured. Total blood cholesterol level, triglyceride, LDL-cholesterol, and free-cholesterol values were lower in rats that were fed 1. plantarum CK 102 than in those not fed L. plantarum CK 102. This cholesterol lowering effect implies that L. plantarum CK 102 could be utilized as an additive for health-assistance foods. In conclusion, these results suggest that the 1. plantarum CK 102 isolated could be used commercially as a probiotic.
Comparison of Hydrogenases from Clostridium butyricum and Thiocapsa roseopersicina: Hydrogenases of C. butyricum and T. roseopersicina
Baek Jin-Sook ; Choi Eun-Hye ; Yun Young-Su ; Kim Sun-Chang ; Kim Mi-Sun ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1210~1215
The properties related to the temperature and oxygen stability of the cytoplasmic hydrogenases from the fermentative strict anaerobic bacterium, Clostridium butyricum NCIB 9576 (Cl. butyricum), and purple sulfur phototrophic bacterium, Thiocapsa roseopersicina NCIB 8347 (T. roseopersicina), were compared. The optimum temperatures for the growth of Cl. butyricum and T. roseopersicina were 37
, respectively, whereas those for the H
evolution of the cytoplasmic hydrogenases prepared from Cl. butyricum (C-H
ase) and T. roseopersicina (T-H
ase) were 45
, respectively. The T-H
ase was more thermostable than the C-H
ase and retained its full activity for 5 h at 50
under anaerobic conditions and 90% of its activity at 60
, whereas the C-H
ase lost its activity drastically at 50
. The optimum pHs for H
oxidation of the C-H
ase and T-H
ase were 9.0 and 7.5, respectively. Both enzymes showed a maximum H
evolution activity at pH 7.0. Under aerobic conditions, 80% of the T-H
ase activity was retained for 10 h at 30
, and 50% of the activity remained after 6 days under the same experimental conditions. However, the C-H
ase was labile to oxygen and lost its activity immediately on exposure to air. Therefore, these properties of the T-H
ase are expected to be advantageous for application in in vitro biological H
Protein Array Fabricated by Microcontact Printing for Miniaturized Immunoassay
Lee Woo-Chang ; Lim Sang-Soo ; Choi Bum-Kyoo ; Choi Jeong-Woo ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1216~1221
A protein array was fabricated for a miniaturized immunoassay using microcontact printing (
CP). A polydimethylsiloxane (PDMS) stamp with a 5
m dimension was molded from a silicon master developed by photolithography. Under optimal fabrication conditions, including the baking, incubation, and exposure time, a silicon master was successfully fabricated with a definite aspect ratio. An antibody fragment was utilized as the ink for the
CP, and transferred to an Au substrate because of the Au-thiol (-SH) interaction. The immobilization and antibody-antigen interaction were investigated with fluorescence microscopy. When human serum albumin (HSA) was applied to the protein array fabricated with an antibody against HSA, the detection limit was 100 pg/ml of HSA when using a secondary antibody labeled with a fluorescence tag. The fabricated protein array maintained its activity for 14 days.
Proteomic Analysis of Protein Expression Patterns Associated with Astaxanthin Accumulation by Green Alga Haematococcus pluvialis (Chlorophyceae) Under High Light Stress
Kim Jeong-Dong ; Lee Woo-Sung ; Kim Beob-Min ; Lee Choul-Gyun ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1222~1228
Two kinds of Haematococcus pluvialis cells (green vegetative cells cultivated under optimal cell culture conditions and red cyst cells maintained under high light stress conditions to induce astaxanthin production) were used to investigate the protein expression profiles by two-dimensional electrophoresis, image analysis, and peptide mass fingerprinting. The cellular accumulation of astaxanthin was evident after exposure to high light intensity and reached the maximum cellular level after 78 h of high light stress. In a 2-D electrophoresis analysis, 22 proteins were upregulated over 2-fold in the red cyst cells when compared with the green vegetative cells and selected for further analysis by chemically assisted fragmentation (CAF)-MALDI-TOF sequencing to identify the protein functions. Among 22 different spots, several key enzymes specific to the carotenoid pathway, including isopentenyl pyrophosphate isomerase (IPP) and lycopene
-cyclase, appeared in H. pluvialis after exposure to high light intensity. Therefore, IPP and lycopene
-cyclase would appear to be involved with carotenoid accumulation in the cytoplasm, as these peptides were preferentially upregulated by high light intensity preceding an increase in carotenoid, and only these forms were detected in the red cyst cells.
Lab-on-a-Chip for Monitoring the Quality of Raw Milk
Choi Jeong-Woo ; Kim Young-Kee ; Kim Hee-Joo ; Lee Woo-Chang ; Seong Gi-Hun ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1229~1235
A lab-on-a-chip (LoC) was designed for simultaneous monitoring of microorganisms, antibiotic residues, somatic cells, and pH in raw milk. The LoC was fabricated from polydimethylsiloxane (PDMS) using microelectromechanical system (MEMS) technology, which consisted of two parts; a protein array and microchannel. The protein array was fabricated by immobilizing five types of antibodies corresponding to two microorganisms, two antibiotic residues, and somatic cells. A sol-gel film was deposited on a glass substrate to immobilize the antibodies. The target analytes in raw milk could be bound with the corresponding antibody by an immunoreaction, and the antigen-antibody complex was detected using fluorescence microscopy. SNARF-dextran was used as a pH indicator, and the SNARF-entrapped hydrogel was attached to the microchannel in the chip. After injecting the milk sample into the channel, the pH was measured by monitoring the change in fluorescence intensity by fluorescence microscopy. The on-chip simultaneous assay of two microorganisms (E. coli O157:H7 and Streptococcus agalactiae), two antibiotic residues (penicillin G and dihydrostreptomycin), and neutrophils was successfully accomplished using the proposed LoC system.
Allyl Alcohol Found in Heated Garlic is a Potent Selective Inhibitor of Yeasts
Lee Se-Hi ; Woo Yong-Ho ; Kyung Kyu-Hang ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1236~1239
Allyl alcohol (2-propen-l-ol), found in considerable amounts in heated garlic, was able to discriminate yeasts from bacteria and was approximately three orders of magnitude more inhibitory towards yeasts than bacteria. The average minimum inhibitory concentration (MIC) of allyl alcohol for bacteria and yeasts was 5.0% and 0.0056%, respectively. The unsaturated primary alcohols, including allyl alcohol and 2-buten-l-ol, seemed to work differently from all the other saturated alcohols and unsaturated secondary alcohols in inhibiting various yeasts. An alcohol dehydrogenase-negative (ADH
) strain of Saccharomyces cerevisiae was as resistant to allyl alcohol as various bacteria, exhibiting an MIC of 5.0%. The unsaturated primary alcohols were apparently oxidized into the corresponding unsaturated aldehydes before they inhibited the yeasts.
Influence of Agitation Intensity and Aeration Rate on Production of Antioxidative Exopolysaccharides from Submerged Mycelial Culture of Ganoderma resinaceum
Kim Hyun-Mi ; Kim Sang-Woo ; Hwang Hye-Jin ; Park Moon-Ki ; Mahmoud Yehia A.-G. ; Choi Jang-Won ; Yun Jong-Won ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1240~1247
The present study investigated the influence of the aeration rate and agitation intensity on the production of the mycelial biomass and antioxidative exopolysaccharide (EPS) in Ganoderma resinaceum. In submerged cultures with varying agitation speeds and aeration rates in a stirred-tank reactor, the maximum mycelial biomass and maximum EPS concentration were achieved at 50 rpm and 300 rpm, respectively. Under varying aeration rates, the highest amount of mycelial biomass (18.1 g/l) was accumulated at the lowest aeration rate (0.5 vvm) and the maximum EPS production (3.0 g/l) obtained at 1.0 vvm. A compositional analysis revealed that the five different EPSs were protein-bound heteropolysaccharides, consisting of 87.17-89.22% carbohydrates and 10.78-12.83% proteins. The culture conditions had a striking affect on the carbohydrate composition of the EPS, resulting in different antioxidative activities. All the EPSs showed strong scavenging activities against superoxide and 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radicals, whereas no clear trend in antioxidative activity was observed against hydroxyl radicals and lipid peroxides. Although the precise reason for this difference is still unclear, the high glucose moiety of EPS is probably linked to its broad spectrum of antioxidative activity.
Effect of Spirulina platensis and Probiotics as Feed Additives on Growth of Shrimp Fenneropenaeus chinensis
Kim Choong-Jae ; Yoon Sook-Kyung ; Kim Hong-Ik ; Park Yong-Ha ; Oh Hee-Mock ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1248~1254
The effect of Spirulina platens is and probiotics as feed additives on the growth of the shrimp Fenneropenaeus chinensis was investigated in comparison with a control. The shrimp were cultured in rearing tanks in a seawater pond for 35 days from September 1, 2004. As regards the water quality, the probiotic treatment (T2, commercial diet and 3% probiotics) produced a lower TDN (total dissolved nitrogen) and TDP (total dissolved phosphorus), making it effective in water quality improvement. Nonetheless, the phytoplankton flora succeeded from diatoms to cyanobacteria, regardless of the feed additives. Treatment T3, including 3% S. platensis, produced the highest mean body weight, which was 39% higher than that for all the other treatments (P<0.05). Accordingly, it was found that the use of Spirulina and probiotics as feed additives increased the shrimp body weight and improved the water quality, respectively.
Hydrolysis of Agricultural Residues and Kraft Pulps by Xylanolytic Enzymes from Alkaliphilic Bacillus sp. Strain BK
Kaewintajuk Kusuma ; Chon Gil-Hyong ; Lee Jin-Sang ; Kongkiattikajorn Jirasak ; Ratanakhanokchai Khanok ; Kyu Khin Lay ; Lee John-Hwa ; Roh Min-Suk ; Choi Yun-Young ; Park Hyun ; Lee Yun-Sik ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1255~1261
An alkaliphilic bacterium, Bacillus sp. strain BK, was found to produce extracellular cellulase-free xylanolytic enzymes with xylan-binding activity. Since the pellet-bound xylanase is eluted with 2% TEA from the pellet of the culture, they contain a xylan-binding region that is stronger than the xylan-binding xylanase of the extracellular enzyme. The xylanases had a different molecular weight and xylan-binding ability. The enzyme activity of xylanase in the extracellular fraction was 6 times higher than in the pellet-bound enzyme. Among the enzymes, xylanase had the highest enzyme activity. When Bacillus sp. strain BK was grown in pH 10.5 alkaline medium containing xylan as the sole carbon source, the bacterium produced xylanase, arabinofuranosidase, acetyl esterase, and
-xylosidase with specific activities of 1.23, 0.11, 0.06, and 0.04 unit per mg of protein, respectively. However, there was no cellulase activity detected in the crude enzyme preparation. The hydrolysis of agricultural residues and kraft pulps by the xylanolytic enzymes was examined at 50
and pH 7.0. The rate of xylan hydrolysis in com hull was higher than those of sugarcane bagasse, rice straw, com cop, rice husk, and rice bran. In contrast, the rate of xylan hydrolysis in sugarcane pulp was 2.01 and 3.52 times higher than those of eucalyptus and pine pulp, respectively. In conclusion, this enzyme can be used to hydrolyze xylan in agricultural residues and kraft pulps to breach and regenerate paper from recycled environmental resources.
Induction of Apoptosis in the HepG2 Cells by HY53, a Novel Natural Compound Isolated from Bauhinia forficata
Lim Hae-Young ; Lim Yoong-Ho ; Cho Youl-Hee ; Lee Chul-Hoon ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1262~1268
In the search for a novel cytotoxic substance from medicinal plants, HY53 (
; molecular weight 296) was isolated from the leaves of Pata de Vaca (Bauhinia forficata). The growth of the HepG2 cells was inhibited in a dose-dependent manner when treated with 0.07 to 0.40 mM HY53 for 24 h (IC
: 0.13 mM). Furthermore, nuclear DAPI staining revealed the typical nuclear features of apoptosis in the HepG2 cells exposed to 0.27 mM HY53, whereas a flow cytometric analysis of the HepG2 cells using propidium iodide showed that the apoptotic cell population increased gradually from 8% at 0 mM to 23% at 0.14 mM and 45% at 0.40 mM after being exposed to each concentration of HY53 for 24 h. Moreover, a TUNEL assay also exhibited the apoptotic induction of the HepG2 cells treated with HY53. To obtain further information on the HY53-induced apoptosis, the expression level of certain apoptosis-associated proteins was examined using a Western blot analysis. Treatment of the HepG2 cells with HY53 resulted in the activation of caspase-3, and subsequent proteolytic cleavage of poly(ADP-ribose) polymerase (PARP). Consequently, the results confirmed that the apoptosis in the HepG2 cells was induced by HY53 and the involvement of caspase-3-mediated PARP cleavage in the apoptotic process.
Selection of Multienzyme Complex-Producing Bacteria Under Aerobic Cultivation
Pason Patthra ; Chon Gil-Hyong ; Ratanakhanokchai Khanok ; Kyu Khin Lay ; Jhee Ok-Hwa ; Kang Ju-Seop ; Kim Won-Ho ; Choi Kyung-Min ; Park Gil-Soon ; Lee Jin-Sang ; Park Hyun ; Rho Min-Suk ; Lee Yun-Sik ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1269~1275
The selection of multienzyme complex-producing bacteria under aerobic condition was conducted for improving the degradation of lignocellulosic substances. The criteria for selection were cellulase and xylanase enzyme production, the presence of cellulose-binding domains and/or xylan-binding domains in enzymes to bind to insoluble substances, the adhesion of bacterial cells to insoluble substances, and the production of multiple cellulases and xylanases in a form of a high molecular weight complex. Among the six Bacillus strains, isolated from various sources and deposited in our laboratory, Paenibacillus curdlanolyticus B-6 strain was the best producer of cellulase and xylanase enzymes, which have both cellulose-binding factors (CBFs) and xylan-binding factors (XBFs). Moreover, multiple carboxymethyl cellulases (CMCases) and xylanases were produced by the strain B-6. The zymograms analysis showed at least 9 types of xylanases and 6 types of CMCases associated in a protein band of xylanase and cellulase with high molecular weight. These cells also enabled to adhere to both avicel and insoluble xylan, which were analyzed by scanning electron microscopy. The results indicated that the strain B-6 produced the multienzyme complex, which may be cellulosome or xylanosome. Thus, P. curdlanolyticus B-6 was selected to study the role and interaction between the enzymes and their substrates and the cooperation of multiple enzymes to enhance the hydrolysis due to the complex structure for efficient cellulases and xylanases degradation of insoluble polysaccharides.
Antibiotic Resistance and Genetic Diversity of Listeria monocytogenes Isolated from Chicken Carcasses in Korea
Jang Sung-Sik ; Choo Eui-Young ; Han Ki-Seon ; Miyamoto Takahisa ; Heu Sung-Gi ; Ryu Sang-Ryeol ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1276~1284
Listeria monocytogenes is a well-known high-risk foodborne pathogen that grows at refrigeration temperature and is responsible for outbreaks of listeriosis. We report here the incidence of L. monocytogenes in fresh chicken carcasses and present genetic diversity of L. monocytogenes isolates. In this study, 25 g of chicken carcasses from markets in Korea were examined according to the FDA method, and presumptive isolates were confirmed by multiplex PCR assay. L. monocytogenes isolates were analyzed by Pulsed-Field Gel Electrophoresis using restriction enzymes, ApaI and AscI, to obtain strain-specific DNA fragments profiles. Antimicrobial resistance of L. monocytogenes strains against generally used antibiotics (Penicillin G, Kanamycin, Tetracycline, Vancomycin, Cephalothin, Rifampicin, Erythromycin, Ampicillin, Gentamicin, Streptomycin, and Chloramphenicol) were analyzed by NCCLS protocols to examine the presence of antimicrobial resistance in natural L. monocytogenes. Of a total 274 chicken samples, 81 samples (29.6%) were positive for L. monocytogenes. Listeria innocua (50.1%), Listeria welshimeri (6.9%), and Listeria grayi (11.3%) were also detected. PFGE analysis, using restriction enzymes ApaI and AscI, showed 27 pulsotypes of L. monocytogenes. Antimicrobial resistance analysis confirmed the existence of antimicrobial resistance for penicillin G and tetracycline in isolated L. monocytogenes strains.
Regulatory Characteristics of the Vibrio vulnificus putAP Operon Encoding Proline Dehydrogenase and Proline Permease
Lee Jeong-Hyun ; Jeong So-Young ; Choi Sang-Ho ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1285~1291
The proline utilization (put) operon of Vibrio vulnificus consists of the putAP genes encoding a proline dehydrogenase and proline permease. The result of put-lux transcriptional fusion analysis suggests that the V vulnificus putAP operon is not autoregulated by the PutA protein. A putR null mutation decreased proline dehydrogenase activity and the level of the put transcripts, indicating that transcription of putAP is under the positive control of PutR. The deduced amino acid sequence of the putR was similar to those reported from other bacteria with high levels of identity. Chromatin IP and GST pull-down assays revealed that PutR specifically binds to the putAP promoter region in vivo, and interacts with CRP in vitro. Taken together, the results suggested that PutR exerts its effect on putAP expression by directly interacting with CRP bound to the upstream region of P
Stimulation of Platelet-Activating Factor (PAF) Synthesis in Human Intestinal Epithelial Cell Line by Aerolysin from Aeromonas encheleia
Nam In-Young ; Cho Jae-Chang ; Myung Hee-Joon ; Joh Ki-Seong ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1292~1300
Aeromonas encheleia, a potential human intestinal pathogen, was shown to infect a human intestinal epithelial cell line (Caco-2) in a noninvasive manner. The transcriptional profile of the Caco-2 cells after infection with the bacteria revealed an upregulated expression of genes involved in chloride secretion, including that of phospholipase A2 (PLA2) and platelet-activating factor (PAF) acetylhydrolase (PAFAH2). This was also confirmed by a real-time RT-PCR analysis. As expected from PLA2 induction, PAF was produced when the Caco-2 cells were infected with the bacteria, and PAF was also produced when the cells were treated with a bacterial culture supernatant including bacterial extracellular proteins, yet lacking lipopolysaccharides. Bacterial aerolysin was shown to induce the production of PAF
Simultaneous Detection of Yersinia enterocolitica, Staphylococcus aureus, and Shigella spp. in Lettuce Using Multiplex PCR Method
Park Si-Hong ; Kim Hyun-Joong ; Kim Hae-Yeong ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1301~1305
The development of rapid, infallible, and sensitive methods of detecting foodborne pathogens has received much impetus in recent years owing to an increased public awareness of the health hazards. For the rapid and simultaneous detection of these foodborne pathogens, a multiplex PCR method was developed. Yersinia enterocolitica, Staphylococcus aureus, and Shigella spp. are bacteria of concern because of their specific growing condition that enables them to live at low temperatures. In order to detect each pathogenic bacterium, specific primers from Y. enterocolitica, St. aureus, and Sh. flexneri were selected and validated successfully. To apply this method to food stored at low temperature, Y. enterocolitica, St. aureus, and Sh. flexneri were artificially inoculated in lettuce and incubated for enrichment. The multiplex PCR assays were able to simultaneously detect three pathogens, and the presence of three bands was observed at initial inoculation levels of approximately 1
CFU/g in lettuce. Therefore, this method could be used for simultaneous detection of Y. enterocolitica, St. aureus, and Shigella spp. contaminated in lettuce during cultivation, transportation, preservation, and storage.
A New Intermediate in the Degradation of Carbofuran by Sphingomonas sp. Strain SB5
Park Myung-Ryeol ; Lee Sun-Woo ; Han Tae-Ho ; Oh Byung-Tack ; Shim Jae-Han ; Kim In-Seon ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1306~1310
Sphingomonas sp. strain SB5 could degrade carbofuran and carbofuran-7-phenol to a hydrolytic product, 2-hydroxy-3-(3-methlypropan-2-o1)phenol, and several red metabolites. However, the chemical structures of the red metabolites have largely remained unidentified. In this study, we identified the structure of one of the red metabolites as 5-(2-hydroxy-2-methyl-propyl)-2,2-dimethyl- 2,3-dihydro-naphtho[2,3-6]furan-4,6,7,9-tetrone by using mass spectrometric and NMR (
C) analyses. It is suggested that the red metabolite resulted from condensation of some metabolites in the degradation of 2-hydroxy-3-(3-methlypropan-2-o1)phenol, a hydrolytic product derived from carbofuran. To our knowledge, this is the first paper to report a red metabolite in bacterial degradation of the insecticide carbofuran.
Characterization of D-Glucose
-1-Phosphate Uridylyltransferase (VldB) and Glucokinase (VIdC) Involved in Validamycin Biosynthesis of Streptomyces hygroscopicus var. limoneus KCCM 11405
Seo Myung-Ji ; Im Eun-Mi ; Singh Deepak ; Rajkarnikar Arishma ; Kwon Hyung-Jin ; Hyun Chang-Gu ; Suh Joo-Won ; Pyun Yu-Ryang ; Kim Soon-Ok ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1311~1315
Aminocyclitol antibiotic validamycin A, a prime control agent for sheath blight disease of rice plants, is biosynthesized by Streptomyces hygroscopicus var. limoneus. Within the validamycin biosynthetic gene cluster, vldBC forms an operon of vldABC with vidA, the gene encoding 2-epi-5-epi-valiolone synthase. Biochemical studies, employing the recombinant proteins from Escherichia coli, established VldB and VldC as D-glucose
-1-phosphate uridylyltransferase and glucokinase, respectively. This finding substantiates that the validamycin biosynthetic gene cluster harbors genes encoding the enzymes for UDP-glucose formation from glucose. Therefore, we propose that validamycin biosynthesis employs its own catalysts to generate UDP-glucose, but not depending on the primary metabolism.
Use of Antibody Displayed Phage for the Detection of Dextran Using a Dipstick Assay and Transmission Electron Micrograph
Kim Du-Woon ; Day Donal F. ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1316~1319
An antibody displayed phage collection (SBAE-2R), screened from a human synthetic phage antibody library (Fab 21ox), was used for the determination of dextran. The dextran-binding affinity was determined by serologically specific transmission electron microscopy (TEM) and a paper dipstick assay. The phage collection was distributed over the dextrancoated grids with 39
on the grids. Phages were not seen on dextran-coated grids exposed to the Fab 2lox phage library. The phage collection (SBAE-2R) produced 54
3 color normalized intensity (N.I.) from 125 ppm to 1,000 ppm of dextran and 5
1 (N.I.) for 63 ppm of dextran in a paper dipstick assay. This research extends the analytical options for dextran analysis by antibody displayed phage with a minimum of equipment usage.
Development of an Agar Diffusion Method to Measure Elastase Inhibition Activity Using Elastin-Congo Red
Jung Kyung-Hwan ; Kim Hyun-Joo ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1320~1324
The pancreatic and neutrophil elastases are associated with several illnesses including lung and vascular diseases, various cancers, and pancreatitis. The development of a potent and specific inhibitor to the elastases could lead to new therapies. In this study, an agar diffusion method was modified to include a substrate-dye conjugate (Elastin-Congo red) as a substrate of elastase and an indicator of elastase inhibitory activity. The Elastin-Congo red agar plates consisted of 0.1 % Elastin-Congo red and 2.5% agar. The elastase and elastase inhibitors were simultaneously loaded into wells, ultimately resulting in halo formations in which the halo diameter decreased as the concentration of elastase inhibitor increased. The concentration of elastase inhibitor in the samples, therefore, was inversely proportional to the halo diameters. This simplified method provided an excellent correlation with the standard microplate technique, which uses a chromogenic substrate. The concentration of elastase inhibitor obtained from the culture supernatant of a recombinant elastase inhibitor produced by the yeast Pichia pastoris was easily determined. This study has established a simple modified and inexpensive agar diffusion method that is potentially useful for the identification, quantification, and screening of new elastase inhibitors.
Substitutions for Cys-472 and His-509 at the Active Site of
-Galactosidase from Lactococcus lactis ssp. lactis 7962 Cause Large Decreases in Enzyme Activity
Chung Hye-Young ; Yang Eun-Ju ; Chang Hae-Choon ;
Journal of Microbiology and Biotechnology, volume 16, issue 8, 2006, Pages 1325~1329
Structural modeling of
-galactosidase from L. lactis ssp. lactis 7962 has shown that the residues Cys-472 and His-509 are located in the wall of the active-site cavity. To examine the functions of Cys-472 and His-509, we generated five site-specific mutants: Cys-472-Ser, Cys-472-Thr, Cys-472-Met, His-509-Asn, and His-509-Phe.
-Galactosidase substituted at Cys-472 with Met or His-509 with Phe had <3% of the activity of the native enzyme when assayed using ONPG as substrate. The other mutants Cys-472-Ser, Cys-472-Thr, and His-509-Asn had ca. 10-15% of the native enzyme activity. The V
values of the five mutated enzymes were lower (60-7,000-fold) than that of native enzyme. These results show that the catalytic ability of
-galactosidase is significantly affected by mutations at Cys-472 or His-509.