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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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The Korean Society for Applied Microbiology and Biotechnology
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Volume & Issues
Volume 18, Issue 12 - Dec 2008
Volume 18, Issue 11 - Nov 2008
Volume 18, Issue 10 - Oct 2008
Volume 18, Issue 9 - Sep 2008
Volume 18, Issue 8 - Aug 2008
Volume 18, Issue 7 - Jul 2008
Volume 18, Issue 6 - Jun 2008
Volume 18, Issue 5 - May 2008
Volume 18, Issue 4 - Apr 2008
Volume 18, Issue 3 - Mar 2008
Volume 18, Issue 2 - Feb 2008
Volume 18, Issue 1 - Jan 2008
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Molecular Diversity of Bacterial Communities from Subseafloor Rock Samples in a Deep-Water Production Basin in Brazil
Von Der Weid, Irene ; Korenblum, Elisa ; Jurelevicius, Diogo ; Rosado, Alexandre Soares ; Dino, Rodolfo ; Sebastian, Gina Vasquez ; Seldin, Lucy ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 5~14
The deep subseafloor rock in oil reservoirs represents a unique environment in which a high oil contamination and a very low biomass can be observed. Sampling this environment has been a challenge owing to the techniques used for drilling and coring. In this study, the facilities developed by the Brazilian oil company PETROBRAS for accessing deep subsurface oil reservoirs were used to obtain rock samples at 2,822-2,828 m below the ocean floor surface from a virgin field located in the Atlantic Ocean, Rio de Janeiro. To address the bacterial diversity of these rock samples, PCR amplicons were obtained using the DNA from four core sections and universal primers for 16S rRNA and for APS reductase (aps) genes. Clone libraries were generated from these PCR fragments and 87 clones were sequenced. The phylogenetic analyses of the 16S rDNA clone libraries showed a wide distribution of types in the domain bacteria in the four core samples, and the majority of the clones were identified as belonging to Betaproteobacteria. The sulfate-reducing bacteria community could only be amplified by PCR in one sample, and all clones were identified as belonging to Gammaproteobacteria. For the first time, the bacterial community was assessed in such a deep subsurface environment.
Characterization of Mutations in AlHK1 Gene from Alternaria longipes: Implication of Limited Function of Two-Component Histidine Kinase on Conferring Dicarboximide Resistance
Luo, Yiyong ; Yang, Jinkui ; Zhu, Mingliang ; Yan, Jinping ; Mo, Minghe ; Zhang, Keqin ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 15~22
Four series (S, M, R, and W) of Alternaria longipes isolates were obtained based on consecutive selection with Dimethachlon (Dim) and ultraviolet irradiation. These isolates were then characterized according to their tolerance to Dim, sensitivity to osmotic stress, and phenotypic properties. All the selected Dim-resistant isolates showed a higher osmosensitivity than the parental strains, and the last generation was more resistant than the first generation in the M, R, and W series. In addition, the changes in the Dim resistance and osmotic sensitivity were not found to be directly correlated, and no distinct morphologic characteristics were found among the resistant and sensitive isolates, with the exception of the resistant isolate K-11. Thus, to investigate the molecular basis of the fungicide resistance, a group III two-component histidine kinase (HK) gene, AlHK1, was cloned from nineteen A. longipes isolates. AlHK1p was found to be comprised of a six 92-amino-acid repeat domain (AARD), HK domain, and response regulator domain, similar to the Os-1p from Neurospora crassa. A comparison of the nucleotide sequences of the AlHK1 gene from the Dim-sensitive and -resistant isolates revealed that all the resistant isolates contained a single-point mutation in the AARD of AlHK1p, with the exception of isolate K-11, where the AlHK1p contained a deletion of 107 amino acids. Moreover, the AlHK1p mutations in the isolates of each respective series involved the same amino acid substitution at the same site, although the resistance levels differed significantly in each series. Therefore, these findings suggested that a mutation in the AARD of AlHK1p was not the sole factor responsible for A. longipes resistance to dicarboximide fungicides.
Evidence on the Presence of
Group I Intron in the Marine Cyanobacterium Synechococcus elongatus
Muralitharan, Gangatharan ; Thajuddin, Nooruddin ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 23~27
Self-splicing group I introns in tRNA anticodon loops have been found in diverse groups of bacteria. In this work, we identified
group I introns in six strains of marine Synechococcus elongatus. Introns with sizes around 280 bp were consistently obtained in all the strains tested. In a phylogenetic analysis using the nucleotide sequence determined in this study with other cyanobacterial
intron sequences, the Synechococcus sequence was grouped together with the sequences from other unicellular cyanobacterial strains. Interestingly, the phylogenetic tree inferred from the intronic sequences clearly separates the different tRNA introns, suggesting that each family has its own evolutionary history.
Molecular Characterization of Regulatory Genes Associated with Biofilm Variation in a Staphylococcus aureus Strain
Kim, Jong-Hyun ; Kim, Cheorl-Ho ; Hacker, Jorg ; Ziebuhr, Wilma ; Lee, Bok-Kwon ; Cho, Seung-Hak ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 28~34
Biofilm formation in association with the intercellular adhesion (icaADBC) gene cluster is a serious problem in nosocomial infections of Staphylococcus aureus. In all 112 S. aureus strains tested, the ica genes were present, and none of these strains formed biofilms. The biofilm formation is known to be changeable by environmental factors. We have found about 30% of phase variation in these strains with treatment of tetracycline, pristinamycin, and natrium chloride. However, this phenotype disappeared without these substances. Therefore, we have constructed stable biofilm-producing variants through a passage culture method. To explain the mechanism of this variation, nucleotide changes of ica genes were tested in strain S. aureus 483 and the biofilm-producing variants. No differences of DNA sequence in ica genes were found between the strains. Additionally, molecular analysis of three regulatory genes, the accessory gene regulator (agr) and the staphylococcal accessory regulator (sarA), and in addition, alternative transcription factor
(sigB), was performed. The data of Northern blot and complementation showed that SigB plays an important role for this biofilm variation in S. aureus 483 and the biofilm-producing variants. Sequence analysis of the sigB operon indicated three point mutations in the rsbU gene, especially in the stop codon, and two point mutations in the rsbW gene. This study shows that this variation of biofilm formation in S. aureus is deduced by the role of sigB, not agr and sarA.
Nonribosomal Peptide Synthase is Responsible for the Biosynthesis of Siderophore in Vibrio vulnificus MO6-24/O
Kim, In-Hwang ; Shim, Jung-Im ; Lee, Ko-Eun ; Hwang, Won ; Kim, Ik-Jung ; Choi, Sang-Ho ; Kim, Kun-Soo ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 35~42
Vibrio vulnificus produces siderophores, low-molecular-weight iron-chelating compounds, to obtain iron under conditions of iron deprivation. To identify genes associated with the biosynthesis of siderophore in V. vulnificus MO6-24/O, we screened clones with mini-Tn5 random insertions for those showing decreased production of siderophore. Among 6,000 clones screened, nine such clones were selected. These clones contain the transposon inserted in VV2_0830 (GenBank accession number) that is a homolog of a nonribosomal peptide synthase (NRPS). There is an another NRPS module, VV2_0831, 49-bp upstream to VV2_0830. We named these two genes vvs (Vibrio vulnificus siderophore synthase) A and B, respectively. Mutation of either vvsA or vvsB showed a decreased production of siderophore. The expression of an NRPS-lux fusion was negatively modulated by the presence of iron, and the regulation was dependent on Fur (ferric uptake regulator). However, the expression of the NRPS genes was still not fully derepressed in the iron-rich condition, even in furnull mutant cells, suggesting that some other unknown factors are involved in the regulation of the genes. We also demonstrated that the NRPS genes are important for virulence of the pathogen in a mice model.
Expression, Purification, Crystallization and Preliminary X-Ray Crystallographic Analysis of CnrX from Cupriavidus metallidurans CH34
Kim, Kook-Han ; Jung, Eun-Jung ; Im, Ha-Na ; Lelie, Daniel Van Der ; Kim, Eunice Eun-Kyeong ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 43~47
The nickel and cobalt resistance of Cupriavidus metallidurans CH34 is mediated by the CnrCBA efflux pump encoded by the cnrYHXCBAT metal resistance determinant. The products of the three genes cnrYXH transcriptionally regulate expression of cnr. CnrY and CnrX are membrane-bound proteins, probably functioning as anti-sigma factors, whereas CnrH is a cnr-specific extracytoplasmic functions (ECF) sigma factor. The periplasmic domain of CnrX (residues 29-148) was cloned as a N-terminal His-tagged protein, expressed in Escherichia coli, and purified using affinity chromatography and gel filtration. The molecular mass was estimated to be about 13.6kDa by size exclusion chromatography, corresponding to a monomer. The tetragonal bipyramid crystals were obtained by mixing an equal volume of protein in 50mM Tris-HCl, pH 7.5, 1% glycerol, 100mM NaCl, 1mM DTT, and the reservoir solution of 15% w/v PEG 2000, 100mM lithium chloride at 277K in 2-4 days using hanging drop vapor diffusion. The protein concentration was 24mg/ml. The crystal that diffracted to
resolution belongs to space group
with unit cell parameters of
, with one molecule of CnrX in the asymmetric unit.
-Aminotransferase from Caulobacter crescentus and Sitedirected Mutagenesis to Broaden Substrate Specificity
Hwang, Bum-Yeol ; Ko, Seung-Hyun ; Park, Hyung-Yeon ; Seo, Joo-Hyun ; Lee, Bon-Su ; Kim, Byung-Gee ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 48~54
-aminotransferase gene, cc3143 (aptA), from Caulobacter crescentus was screened by bioinformatical tools and overexpressed in E. coli, and the substrate specificity of the
-aminotransferase was investigated. AptA showed high activity for short-chain
-amino acids. It showed the highest activity for 3-amino-n-butyric acid. It showed higher activity toward aromatic amines than aliphatic amines. The 3D model of the
-aminotransferase was constructed by homology modeling using a dialkylglycine decarboxylase (PDB ID: 1DGE) as a template. Then, the
-aminotransferase was rationally redesigned to increase the activity for 3-amino-3-phenylpropionic acid. The mutants N285A and V227G increased the relative activity for 3-amino-3-phenylpropionic acid to 3-amino-n-butyric acid by 11-fold and 3-fold, respectively, over that of wild type.
Overexpression, Crystallization, and Preliminary X-Ray Crystallographic Analysis of the Alanine Racemase from Enterococcus faecalis v583
Priyadarshi, Amit ; Lee, Eun-Hye ; Sung, Min-Woo ; Kim, Jae-Hee ; Ku, Min-Je ; Kim, Eunice Eun-Kyeong ; Hwang, Kwang-Yeon ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 55~58
Alanine racemase, a bacterial enzyme belonging to the fold-type III group of pyridoxal 5'-phosphate (PLP)-dependent enzymes, has been shown to catalyze the interconversion between L- and D-alanine. The alanine racemase from the pathogenic bacterium Enterococcus faecalis v583 has been overexpressed in E. coli and was shown to crystallize an enzyme at 295 K, using polyethylene glycol (PEG) 8000 as a precipitant. X-ray diffraction data to
has been collected using synchrotron radiation. The crystal is a member of the orthorhombic space group,
with unit cell parameter of a=94.634, b=156.516,
. Two or three monomers are likely to be present in the asymmetric unit, with a corresponding
and a solvent content of 63.7% and 45.5%, respectively.
Overexpression, Purification, and Preliminary X-Ray Crystallographic Studies of Methionine Sulfoxide Reductase B from Bacillus subtilis
Park, Ae-Kyung ; Shin, Youn-Jae ; Moon, Jin-Ho ; Kim, Young-Kwan ; Hwang, Kwang-Yeon ; Chi, Young-Min ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 59~62
The peptide methionine sulfoxide reductases (Msrs) are enzymes that catalyze the reduction of methionine sulfoxide back to methionine. Because of two enantiomers of methionine sulfoxide (S and R forms), this reduction reaction is carried out by two structurally unrelated classes of enzymes, MsrA (E.C. 188.8.131.52) and MsrB (E.C. 184.108.40.206). Whereas MsrA has been well characterized structurally and functionally, little information on MsrB is available. The recombinant MsrB from Bacillus subtilis has been purified and crystallized by the hanging-drop vapor-diffusion method, and the functional and structural features of MsrB have been elucidated. The crystals belong to the trigonal space group P3, with unit-cell parameters a=b=136.096,
, and diffracted to
resolution using a synchrotron-radiation source at Pohang Light Source. The asymmetric unit contains six subunits of MsrB with a crystal volume per protein mass
and a solvent content of 63.5%.
Effect of Rhamnolipids on Degradation of Anthracene by Two Newly Isolated Strains, Sphingomonas sp. 12A and Pseudomonas sp. 12B
Cui, Chang-Zheng ; Zeng, Chi ; Wan, Xia ; Chen, Dong ; Zhang, Jia-Yao ; Shen, Ping ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 63~66
Anthracene is a PAH that is not readily degraded, plus its degradation mechanism is still not clear. Thus, two strains of anthracene-degrading bacteria were isolated from long-term petroleum-polluted soil and identified as Sphingomonas sp. 12A and Pseudomonas sp. 12B by a 16S rRNA sequence analysis. To further enhance the anthracene-degrading ability of the two strains, the biosurfactants produced by Pseudomonas aeruginosa
were used, which were characterized as rhamnolipids. It was found that these rhamnolipids dramatically increased the solubility of anthracene, and a reverse-phase HPLC assay showed that the anthracene degradation percentage after 18 days with Pseudomonas sp. 12B was significantly enhanced from 34% to 52%. Interestingly, their effect on the degradation by Sphingomonas sp. 12A was much less, from 35% to 39%. Further study revealed that Sphingomonas sp. 12A also degraded the rhamnolipids, which may have hampered the effect of the rhamnolipids on the anthracene degradation.
Antiviral Activity of the Exopolysaccharide Produced by Serratia sp. Strain Gsm01 Against Cucumber Mosaic Virus
Ipper, Nagesh S. ; Cho, Sae-Youll ; Lee, Seon-Hwa ; Cho, Jun-Mo ; Hur, Jang-Hyun ; Lim, Chun-Keun ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 67~73
The potential of the exopolysaccharide (EPS) from a Serratia sp. strain Gsm01 as an antiviral agent against a yellow strain of Cucumber mosaic virus (CMV-Y) was evaluated in tobacco plants (Nicotiana tabacum cv. Xanthi-nc). The spray treatment of plants using an EPS preparation, 72h before CMV-Y inoculation, protected them against symptom appearance. Fifteen days after challenge inoculation with CMV-Y, 33.33% of plants showed mosaic symptoms in EPS-treated plants compared with 100% in the control plants. The EPS-treated plants, which showed mosaic symptoms, appeared three days later than the controls. The enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR) analyses of the leaves of the protected plants revealed that the EPS treatment affected virus accumulation in those plants. Analysis of phenylalanine ammonia lyase, peroxidase, and phenols in protected plants revealed enhanced accumulation of these substances. The pathogenesis-related (PR) genes expression represented by PR-lb was increased in EPS-treated plants. This is the first report of a systemic induction of protection triggered by EPS produced by Serratia sp. against CMV-Y.
Chemical Composition and Biological Activities of Essential Oils Extracted from Korean Endemic Citrus Species
Baik, Jong-Seok ; Kim, Sang-Suk ; Lee, Jung-A ; Oh, Tae-Heon ; Kim, Ji-Young ; Lee, Nam-Ho ; Hyun, Chang-Gu ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 74~79
The aim of this study was to analyze the chemical composition of 14 kinds of citrus oils and to test their biological activities. Citrus essential oils were obtained by steam distillation from immature fruits collected from Jeju Island and were analyzed using gas chromatograph (GC)-flame ionization detectors (FID) and GC-MS. Limonene (55.4% to 91.7%), myrcene (2.1% to 32.1%),
-pinene (0.6% to 1.6%) and linalool (0.4% to 6.9%) were the major components in most citrus species. To evaluate in vitro antibacterial activity, all essential oils were tested against Propionibacterium acnes and Staphylococcus epidermidis. Nine out of fourteen citrus oils exhibited antibacterial activity against P. acnes, but not against S. epidermidis. The effects of the citrus oils on DPPH radical scavenging, superoxide radical anion scavenging, nitric oxide radical, and cytotoxicity were also assessed. Three essential citrus oils, Joadeung, Dongjunggyul, and Bujiwha, exhibited potent inhibitory effects on nitric oxide production. Two essential oils, Dongjunggyul and Joadeung, showed potent free radical scavenging activities in the DPPH assay. For future applications in cosmetic products, we also performed MTT assays in a human dermal fibroblast cell line. The majority of the essential oils showed no cytotoxicity. The results indicate that citrus essential oils can be useful natural agents for cosmetic application.
Chitosan Oligosaccharides Inhibit Adipogenesis in 3T3-L1 Adipocytes
Cho, Eun-Jae ; Rahman, Atiar ; Kim, Sang-Woo ; Baek, Yu-Mi ; Hwang, Hye-Jin ; Oh, Jung-Young ; Hwang, Hee-Sun ; Lee, Sung-Hak ; Yun, Jong-Won ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 80~87
The 3T3-L1 cell line is a well-established and commonly used in vitro model to assess adipocyte differentiation. Over the course of several days, confluent 3T3-L1 cells can be converted to adipocytes in the presence of an adipogenic cocktail. In this study, the effects of chitosan oligosaccharides (CO) on adipocyte differentiation of 3T3-L1 cells were studied. The CO significantly decreased lipid accumulation, a marker of adipogenesis, in a dose-dependent manner. The low molecular mass CO (1-3 kDa) were the most effective at inhibiting adipocyte differentiation. Moreover, mRNA expression levels of both CCAAT/enhancer-binding protein (C/EBP)
and peroxisome proliferator-activated receptor (PPAR)
, the key adipogenic transcription factors, were markedly decreased by CO treatments. CO also significantly down regulated adipogenic marker proteins such as leptin, adiponectin, and resistin. Our results suggest a role for CO as antiobesity agents by inhibiting adipocyte differentiation mediated through the down regulated expression of adipogenic transcription factors and other specific genes.
Genetically Engineered Biosynthesis of Macrolide Derivatives Including 4-Amino-4,6-Dideoxy-L-Glucose from Streptomyces venezuelae YJ003-OTBP3
Pageni, Binod Babu ; Oh, Tae-Jin ; Liou, Kwang-Kyoung ; Yoon, Yeo-Joon ; Sohng, Jae-Kyung ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 88~94
Two sugar biosynthetic cassette plasm ids were used to direct the biosynthesis of a deoxyaminosugar. The pOTBP1 plasmid containing TDP-glucose synthase (desIII), TDP-glucose-4,6-dehydratase (desIV), and glycosyltransferase (desVII/desVIII) was constructed and transformed into S. venezuelae YJ003, a strain in which the entire gene cluster of desosamine biosynthesis is deleted. The expression plasmid pOTBP3 containing 4-aminotransferase (gerB) and 3,5-epimerase (orf9) was transformed again into S. venezuelae YJ003-OTBP1 to obtain S. venezuelae YJ003-OTBP3 for the production of 4-amino-4,6-dideoxy-L-glucose derivatives. The crude extracts obtained from S. venezuelae ATCC 15439, S. venezuelae YJ003, and S. venezuelae YJ003-OTBP3 were further analyzed by TLC, bioassay, HPLC, ESI/MS, LC/MS, and MS/MS. The results of our study clearly shows that S. venezuelae YJ003-OTBP3 constructs other new hybrid macrolide derivatives including 4-amino-4,6-dideoxy-L-glycosylated YC-17 (3, [M+
] m/z=464.5), methymycin (4, m/z=480.5), novamethymycin (6, m/z=496.5), and pikromycin (5, m/z=536.5) from a 12-membered ring aglycon (10-deoxymethynolide, 1) and a 14-membered ring aglycon (narbonolide, 2). These results suggest a successful engineering of a deoxysugar pathway to generate novel hybrid macrolide derivatives, including deoxyaminosugar.
Immunostimulatory Activities of Polysaccharides from Liquid Culture of Pine-Mushroom Tricholoma matsutake
Kim, Joo-Young ; Byeon, Se-Eun ; Lee, Yong-Gyu ; Lee, Ji-Yeon ; Park, Jong-Sun ; Hong, Eock-Ki ; Cho, Jae-Youl ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 95~103
Mushrooms are regarded as one of the well-known foods and biopharmaceutical materials with a great deal of interest. Polysaccharide
-glucan is the major component of mushrooms that displays various biological activities such as antidiabetic, anticancer, and antihyperlipidemic effects. In this study, we compared the immunostimulatory potency of polysaccharide fractions, prepared from liquid culture of pine-mushroom Tricholoma matsutake, with a potent immunogen lipopolysaccharide (LPS), and their molecular mechanisms on the functional activation of macrophages. We found that fraction II (TMF-II) was able to comparably upregulate or highly enhance the phenotypic functions of macrophages such NO production and cytokine (IL-
, IL-6, IL-12, and TNF-
) expression, to LPS. TMF-II triggered the phosphorylation of
, a critical step for NF-
activation and translocation. Of the upstream signaling enzymes tested, Src and Akt were thought to be the responsible upstream signaling components in induction of NO production, although TMF-II strongly upregulated the phosphorylation of all MAPK pathways. Therefore, our data suggest that T. matsutake-derived
-glucan may exert its immunostimulating activities with similar potency to LPS via activation of multiple signaling pathways linked to NF-
Morphological Variation of Enterobacter sp. BL-2 in Acetate-mediated pH Environment for Excretive Production of Cationic Microbial Polyglucosamine Biopolymer
Son, Mi-Kyung ; Hong, Soo-Jung ; SaGong, Kuk-Hwa ; Lee, Yong-Hyun ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 104~106
Enterobacter sp. BL-2 excretively produced a unique cationic polyglucosamine biopolymer PGB-1 comprised of more than 95% D-glucosamine in an acetate-mediated culture condition. The excretion of the biopolymer PGB-1 was closely associated with the cellular morphology of Enterobacter sp. BL-2, a feature highly dependable on the pH of the medium. The initially formed uneven and irregular surface cells were aggregated into the cell-biopolymer network structure connected by the adhesion modules of the cell-bound biopolymer. The excretive production of the biopolymer PGB-1 coincided with the disruption of the cell-biopolymer network, most actively at the medium pH of 8.0.
Peroxidase-mediated Formation of the Fungal Polyphenol 3,14'-Bihispidinyl
Lee, In-Kyoung ; Yun, Bong-Sik ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 107~109
Medicinal fungi, Phellinus linteus and Inonotus xeranticus, produce a cluster of yellow pigment in their fermentation broth that acts as an important element of biological activity. The pigment is composed of diverse polyphenols with a styrylpyrone moiety, mainly hispidin and its dimers, 3,14'-bihispidinyl, hypholomine B, and 1,1-distyrylpyrylethan. Although dimeric hispidins were proposed to be biosynthesized from two molecules of monomer via oxidative coupling by ligninolytic enzymes, laccase and peroxidase, the details of this process remain unknown. In this preliminary study, we attempted to achieve enzymatic synthesis of the hispidin dimer from hispidin by using commercially available horseradish peroxidase (HRP). Consequently, a hispidin dimer, 3,14'-bihispidinyl, was synthesized, whereas the other dimers, hypholomine B and 1,1-distyrylpyrylethan, were not produced. This result suggested that the oxidative coupling at the C-3 and C-14' positions of hispidins was dominant in the process of dimerization by HRP, and indicated that additional catalysts or substrates would be needed to synthesize other hispidin dimers present in the fungal metabolite.
Statistically Designed Enzymatic Hydrolysis for Optimized Production of Icariside II as a Novel Melanogenesis Inhibitor
Park, Jun-Seong ; Park, Hye-Yoon ; Rho, Ho-Sik ; Ahn, Soo-Mi ; Kim, Duck-Hee ; Chang, Ih-Seop ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 110~117
Three kinds of prenylated flavonols, icariside I, icariside II, and icaritin, were isolated from an icariin hydrolysate and their effects on melanogenesis evaluated based on mushroom tyrosinase inhibition and quantifying the melanin contents in melanocytes. Although none of the compounds had an effect on tyrosinase activity, icariside II and icaritin both effectively inhibited the melanin contents with an
of 10.53 and
, respectively. Whereas icariside II was obtained from a reaction with
-glucosidase and cellulase, the icariin was not completely converted into icariside II. Thus, for the high-purity production of icariside II, the reaction was optimized using the response surface methodology, where an enzyme concentration of 5.0mg/ml, pH 7,
, and 8 h reaction time were selected as the central conditions for the central composite design (CCD) for the enzymatic hydrolysis of icariin into icariside II using cellulase. Empirical models were developed to describe the relationships between the operating factors and the response (icariside II yield). A statistical analysis indicated that all four factors had a significant effect (p<0.01) on the icariside II production. The coefficient of determination
was good for the model (0.9853), and the optimum production conditions for icariside II was an enzyme concentration of 7.5mg/ml, pH 5,
, and 12 h reaction time. A good agreement between the predicted and experimental data under the designed optimal conditions confirmed the usefulness of the model. A laboratory pilot scale was also successful.
A New Method for Antimicrobial Susceptibility Testing of Vitro-cultured Bacteria by Means of Resonance Light Scattering Technique
Shi, Yu-Jun ; Chen, Jun ; Xu, Ming ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 118~123
A new method for antimicrobial susceptibility testing of vitro-cultured bacteria on an ordinary fluorescence spectrometer was developed. The viable bacteria reduced 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) to produce insoluble particles that displayed intense resonance scattering light. The assay showed a linear relationship between the number of viable bacteria and the intensity of resonance scattering light. Dead bacteria were unable to reduce MTT. Methicillin-resistant Staphylococcus aureus exposed to flavonoids from Marchantia convoluta showed a flavonoids concentration-dependent inhibition of the ability to reduce MTT. In the assay, less than 12 h was required to attain susceptibility results and fewer bacteria were utilized than in traditional methods. The RLS technique could, in combination with the MTT assay, be a rapid and sensitive measuring method to determine the in vitro activity of new antimicrobials.
Assimilation of Peptides and Amino Acids and Dissimilation of Lactate During Submerged Pure Cultures of Penicillium camembertii and Geotrichum candidum
Aziza, M. ; Adour, L. ; Amrane, A. ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 124~127
The behavior of Penicillium camembertii and Geotrichum candidum growing in submerged pure cultures on simple (glutamate) or complex (peptones) substrates as nitrogen and carbon sources and lactate as a second carbon source was examined. Similar to the behavior previously recorded on a simple substrate (glutamate), a clear differentiation between the carbon source and the energy source was also shown on peptones and lactate during P. camembertii growth, since throughout growth, lactate was only dissimilated, viz., used for energy supply by oxidation into
, whereas peptides and amino acids from peptones were used for carbon (and nitrogen) assimilation. Because of its deaminating activity, G candidum preferred peptides and amino acids to lactate as energy sources, in addition to being assimilated as carbon and nitrogen sources. From this, on peptones and lactate, G candidum grew faster than P. camembertii (0.19 and 0.08 g/l/h, respectively) by assimilating the most readily utilizable peptides and amino acids; however, owing to its lower proteolytic activity, the maximum biomass was lower than that of P. camembertii (3.7 and 5.5 g/l, respectively), for which continuous proteolysis and assimilation of peptides were shown.
Kinetics of Cell Growth and Cyclosporin A Production by Tolypocladium inflatum when Scaling Up from Shake Flask to Bioreactor
El Enshasy, H. ; Fattah, Y. Abdel ; Atta, A. ; Anwar, M. ; Omar, H. ; Magd, S. Abou El ; Zahra, R. Abou ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 128~134
The kinetics of cell growth and Cyclosporin A (Cyc A) production by Tolypocladium inflatum were studied in shake flasks and bioreactors under controlled and uncontrolled pH conditions. In the case of the shake flask, the production time was extended to 226 h and the maximal antibiotic concentration was 76 mg/l. When scaling up the cultivation process to a bioreactor level, the production time was reduced to only 70h with a significant increase in both the cell growth and the antibiotic production. The maximal dry cell weights in the case of the controlled pH and uncontrolled pH cultures in the bioreactor were 22.4g/l and 14.2g/l, respectively. The corresponding maximal dry cell weight values did not exceed 7.25g/l with the shake flask cultures. The maximal values for Cyc A production were 144.72 and 131.4 mg/l for the controlled and uncontrolled pH cultures, respectively. It is also worth noting that a significant reduction was observed in both the dry cell mass and the antibiotic concentration after the Cyc A production phase, whereas the highest rate of antibiotic degradation was observed in the stirred tank bioreactor with an uncontrolled pH. Morphological characterization of the micromorphological cell growth (mycelial/pellet forms) was also performed during cultivation in the bioreactor.
Selective Production of Epothilone B by Heterologous Expression of Propionyl-CoA Synthetase in Sorangium cellulosum
Han, Se-Jong ; Park, Sang-Woo ; Kim, Byung-Woo ; Sim, Sang-Jun ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 135~137
The metabolic engineering of epothilones, as secondary metabolites, was investigated using Sorangium cellulosum to achieve the selective production of epothilone B, a potent anticancer agent. Thus, the propionyl-CoA synthetase gene (prpE) from Ralstonia solanacearum was heterologously expressed in S. cellulosum to increase the production of epothilone B. Propionyl-CoA synthetase converts propionate into propionyl-CoA, a potent precursor of epothilone B. The recombinant S. celluloslim containing the prpE gene exhibited a significant increase in the resolution of epothilones B/A, with an epothilone B to A ratio of 127 to 1, which was 100 times higher than that of the wild-type cells, demonstrating its potential use for the selective production of epothilone B.
Statistical Optimization of Growth Medium for the Production of the Entomopathogenic and Phytotoxic Cyclic Depsipeptide Beauvericin from Fusarium oxysporum KFCC 11363P
Lee, Hee-Seok ; Song, Hyuk-Hwan ; Ahn, Joong-Hoon ; Shin, Cha-Gyun ; Lee, Gung-Pyo ; Lee, Chan ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 138~144
The production of the entomopathogenic and phytotoxic cyclic depsipeptide beauvericin (BEA) was studied in submerged cultures of Fusarium oxysporum KFCC 11363P isolated in Korea. The influences of various factors on mycelia growth and BEA production were examined in both complete and chemically defined culture media. The mycelia growth and BEA production were highest in Fusarium defined medium. The optimal carbon and nitrogen sources for maximizing BEA production were glucose and
, respectively. The carbon/nitrogen ratio for maximal production of BEA was investigated using response surface methodology (RSM). Equations derived by differentiation of the RSM model revealed that the production of BEA was maximal when using 108 mM glucose and 25 mM
Volatile Organic Compound Specific Detection by Electrochemical Signals Using a Cell-Based Sensor
Chung, Sang-Gwi ; Kim, Jo-Chun ; Park, Chong-Ho ; Ahn, Woong-Shick ; Kim, Yong-Wan ; Choi, Jeong-Woo ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 145~152
A cell-based in vitro exposure system was developed to determine whether oxidative stress plays a role in the cytotoxic effects of volatile organic compounds (VOCs) such as benzene, toluene, xylene, and chlorobenzene, using human epithelial HeLa cells. Thin films based on cysteine-terminated synthetic oligopeptides were fabricated for immobilization of the HeLa cells on a gold (Au) substrate. In addition, an immobilized cell-based sensor was applied to the electrochemical detection of the VOCs. Layer formation and immobilization of the cells were investigated with surface plasmon resonance (SPR), cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). The adhered living cells were exposed to VOCs; this caused a change in the SPR angle and the VOC-specific electrochemical signal. In addition, VOC toxicity was found to correlate with the degree of nitric oxide (NO) generation and EIS. The primary reason for the marked increase in impedance was the change of aqueous electrolyte composition as a result of cell responses. The p53 and NF-
downregulation were closely related to the magnitude of growth inhibition associated with increasing concentrations of each VOC. Therefore, the proposed cell immobilization method, using a self-assembly technique and VOC-specific electrochemical signals, can be applied to construct a cell microarray for onsite VOC monitoring.
Adenoviral Vector Mediates High Expression Levels of Human Lactoferrin in the Milk of Rabbits
Han, Zeng-Sheng ; Li, Qing-Wang ; Zhang, Zhi-Ying ; Yu, Yong-Sheng ; Xiao, Bo ; Wu, Shu-Yun ; Jiang, Zhong-Liang ; Zhao, Hong-Wei ; Zhao, Rui ; Li, Jian ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 153~159
The limitations in current technology for generating transgenic animals, such as the time and the expense, hampered its extensive use in recombinant protein production for therapeutic purpose. In this report, we present a simple and less expensive alternative by directly infusing a recombinant adenovirus vector carrying human lactoferrin cDNA into rabbit mammary glands. The milk serum was collected from the infected mammary gland 48 h post-infection and subjected to a 10% SDS-PAGE and Western blotting. An 80-kDa protein was visualized after viral vector infection. With this method, we obtained a high level of expressed human lactoferrin of up to 2.3mg/ml in the milk. Taken together, the method is useful for the transient high-level expression recombinant proteins, and the approach established here is probably one of the most economical and efficient ways for large-scale production of recombinant proteins of biopharmaceutical interest.
Gene Cloning, Expression, and Characterization of a Novel
-Mannanase from Bacillus circulans CGMCC 1416
Li, Yanan ; Yang, Peilong ; Meng, Kun ; Wang, Yaru ; Luo, Huiying ; Wu, Ningfeng ; Fan, Yuliu ; Yao, Bin ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 160~166
A DNA fragment containing 2,079 base pairs from Bacillus circulans CGMCC 1416 was cloned using degenerate PCR and inverse PCR. An open reading frame containing 981 bp was identified that encoding 326 amino acids residues, including a putative signal peptide of 31 residues. The deduced amino acid sequence showed the highest identity (68.1%) with
from Bacillus circulans strain K-1 of the glycoside hydrolase family 5 (GH5). The sequence encoding the mature protein was cloned into the pET-22b(+) vector and expressed in Escherichia coli as a recombinant fusion protein containing an N-terminal hexahistidine sequence. The fusion protein was purified by
affinity chromatography and its hexahistidine tag cleaved to yield a 31-kDa
-mannanase having a specific activity of 481.55U/mg. The optimal activity of the purified protein, MANB48, was at
and pH 7.6. The hydrolysis product on substrate locust bean gum included a monosaccharide and mainly oligosaccharides. The recombinant MANB48 may be of potential use in the feed industry.
Mycoparasitism of Acremonium strictum BCP on Botrytis cinerea, the Gray Mold Pathogen
Choi, Gyung-Ja ; Kim, Jin-Cheol ; Jang, Kyoung-Soo ; Cho, Kwang-Yun ; Kim, Heung-Tae ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 167~170
A fungal strain BCP, which parasitizes Botrytis cinerea gray mold pathogen, was isolated and identified as Acremonium strictum. BCP strain overgrew the colonies of B. cinerea and caused severe lysis of the host hyphae. Frequent penetration and hyphal growth of A. strictum BCP inside the mycelia of B. cinerea were observed under light microscopy. In addition, some morphological abnormalities such as granulation and vacuolation of the cytoplasm were observed in mycelia and spores of B. cinerea. In dual culture test, A. strictum BCP strongly inhibited the mycelial growth of several plant pathogenic fungi as well as B. cinerea. To our knowledge, this is the first report on mycoparasitism of Acremonium species on B. cinerea.
Translocation of VP1686 Upregulates RhoB and Accelerates Phagocytic Activity of Macrophage Through Actin Remodeling
Bhattacharjee, Rabindra N. ; Park, Kwon-Sam ; Chen, Xiuhao ; Iida, Tetsuya ; Honda, Takeshi ; Takeuchi, Osamu ; Akira, Shizuo ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 171~175
Here, we report that Vibrio parahaemolyticus induces a rapid remodeling of macrophage actin and activates RhoB GTPase. Mutational analysis revealed that the effects depend on type III secretion system 1 regulated translocation of a V. parahaemolyticus effector protein, VP1686, into the macrophages. Remodeling of actin is shown to be necessary for increased bacterial uptake followed by initiation of apoptosis in macrophages. This provides evidence for functional association of the VP1686 in triggering an eat me-and-die signal to the host.
Inactivation of S. epidermidis, B. subtilis, and E. coli Bacteria Bioaerosols Deposited on a Filter Utilizing Airborne Silver Nanoparticles
Lee, Byung-Uk ; Yun, Sun-Hwa ; Ji, Jun-Ho ; Bae, Gwi-Nam ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 176~182
In the present study, a control methodology utilizing airborne silver nanoparticles is suggested and tested with respect to its potential to control Gram-positive Staphylococcus epidermidis and Bacillus subtilis, and Gram-negative Escherichia coli bacteria bioaerosols deposited on filters. As it is known that the Gram-negative bacteria are sensitive to airflow exposure, the main focus of this study for testing the airborne silver nanoparticles effect was the Gram-positive Staphylococcus epidermidis and Bacillus subtilis bacteria bioaerosols whereas Escherichia coli bioaerosols were utilized for comparison. Airborne bacteria and airborne silver nanoparticles were quantitatively generated in an experimental system. Bioaerosols deposited on the filter were exposed to airborne silver nanoparticles. The physical and biological properties of the airborne bacteria and airborne silver nanoparticles were measured via aerosol measurement devices. From the experimental results, it was demonstrated that this method utilizing airborne silver nanoparticles offers potential as a bioaerosol control methodology.
Overproduction of Recombinant Human VEGF (Vascular Endothelial Growth Factor) in Chinese Hamster Ovary Cells
Lee, Seong-Baek ; Park, Jeong-Soo ; Lee, Seung-Hee ; Park, Jun-Ho ; Yu, Sung-Ryul ; Kim, Hee-Chan ; Kim, Dong-Jun ; Byun, Tae-Ho ; Baek, Kwang-Hee ; Ahn, Young-Joon ; Yoon, Jae-Seung ;
Journal of Microbiology and Biotechnology, volume 18, issue 1, 2008, Pages 183~187
Vascular endothelial growth factors (VEGFs) are a family of proteins that mediate angiogenesis.
is a VEGF-A isoform and has been extensively studied owing to its potential use in therapeutic angiogenesis. This study established Chinese hamster ovary (CHO) cells overexpressing recombinant human
protein. The production rate of the established CHO cells was over 80mg/l of
protein from a 7-day batch culture process using a 7.5-l bioreactor with a 5-l working volume and serum-free medium. The
protein was purified to homogeneity from the culture supernatant using a two-step chromatographic procedure that resulted in a 48% recovery rate. The purified
protein was a glycosylated homodimeric protein with a higher molecular weight (MW) than the protein expressed from insect cells, suggesting that the glycosylation of the
protein in CHO cells differed from that in insect cells. The purified
protein in this study was functionally active with a half-maximal effective concentration of 3.8ng/ml and specific activity of