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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
> Journal Vol & Issue
Microbiology and Biotechnology Letters
Journal Basic Information
Journal DOI :
The Korean Society for Applied Microbiology and Biotechnology
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Volume & Issues
Volume 26, Issue 6 - Dec 1998
Volume 26, Issue 5 - Oct 1998
Volume 26, Issue 4 - Aug 1998
Volume 26, Issue 3 - Jun 1998
Volume 26, Issue 2 - Apr 1998
Volume 26, Issue 1 - Feb 1998
Selecting the target year
Isolation and Identification of Enterococcus faecalis 2B4-1 Containing Antitumor Substances.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 471~475
The aim of the present research program was to develop a strain of gastrointestinal bacteria containing antitumor substances. Fecal samples were collected from neonates and a number of gastrointestinal bacteria were isolated from the fecal samples by applying selective agar for intestinal bacteria. Among 127 isolates, a strain 2B4-1 containing an antitumor substance against stomach cancer, SNU-1, was selected. The strain 2B4-1 was identified as a strain similar to Enterococcus faecalis NCTC 775 with respect to morphological characteristics, growth temperature, salt and acid tolerance, growth under facultative anaerobic conditions and utilization of carbon sources such as arabinose and melibiose and so on. However, it showed some differences such as a negative reaction to hippurate hydrolysis and negative reaction to
-hemolysis. We assigned to the strain 2B4-1 to Enterococcus faecalis.
Isolation and Characterization of Oxygen-tolerant Mutant of Bifidobacterium longum.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 476~482
Growth sensitivity of bifidobacteria on oxygen hindered their industrial applications so that it was necessary to select oxygen-tolerant strains. Studies on their responses to oxygen might facilitate the effective utilization of bifidobacteria in industry. Oxygen-tolerant strain of Bifidobacterium longum JI-1 was able to remove 3% dissolved oxygen within 10 min whilst oxygen-sensitive strain of B. adolescentis, slime non-former, was not. The ability to remove environmental oxygen seemed to be related to the oxygen-tolerance of bifidobacteria. Mutant B. longum ADJ-1 was induced from the B. longum JI-1 under microaerobic atmosphere. There were no differences in sugar utilization pattern, NADH oxidative enzymes and cellular fatty acid compositions between them. The maximal cell density of the mutant was a little bit reduced to 81% of that of the mother strain. However, the mutant formed thick slime layer around its cell. The layer visualized with confocal scanning laser microscopy from the mutant was 6
in diameter but that from the mother strain was only 3
. Therefore, the improved tolerances of the mutant might come from the slime layer, indicating the increase of the layer might be one of oxygen tolerance mechanisms for bifidobacteria.
Expression, Purification and Characterization of Yeast Thioredoxin System.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 483~489
We carried out the expression and characterization of yeast thioredoxin system including thioredexin 1 (Trx1), Trx2, thioredoxin reductase (TR), and a novel thioredoxin (Trx3), which was reported in the data base of Saccharomyces genome. The Trx1, 2 and TR were expressed as soluble proteins in E. coli and the sizes of purified proteins were equal to the reported their molecular weights. The expressed Trx3 was found in both soluble fraction and precipitate. The size of Trx3 purified from soluble fraction of E. coli crude extracts was estimated as 14 kDa on SDS-PAGE instead of 18 kDa for Trx3 in precipitate. N-terminal amino acid sequence of the small size of purified Trx3 from soluble fraction was analyzed as FQSSYTS which is correspond to the sequence from 20 to 26 for Trx3. Trx3 together with thioredoxin reductase and NADPH was able to reduce the disulfide bridge of insulin and 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB). Trx3 stimulated the antioxidant effect of thioredoxin peroxidase 1 (TPx1) which inhibited inactivation of glutamine synthetase (GS) in dithiothreitol (DTT) containing metal catalyzed oxidation system. The stimulation effect of Trx3 was 10% of the effect of either Trx1 or Trx2. In addition, Trx3 could reduce the disulfide of TPx to thiol, so that the TPx had thioredoxin dependant peroxidase activity. In western blotting analysis, antibodies against purified Trx3 did not cross-react with crude extracts of yeast, purified Trx1, and Trx2 proteins. But, in PCR reaction using the cDNA library of yeast as a template, gene encoding of trx3 was amplified.
Strain Improvement of Aspergillus oryzae for Increasing Productivity of a Proteolytic Enzyme.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 490~496
Aspergillus oryzae producing high proteolytic enzyme was isolated from soybean koji and named tentatively A. oryzae O-1. A. oryzae U-1 was obtained by mutation of A. oryzae O-1 with ultraviolet (UV)-irradiation and produced 14 times higher pretense activity compared with A. oryzae O-1. A. oryzae E-1 was acquired by treatment of A. oryzae U-1 with 0.5 M ethylmethanesulfonate (EMS) for 6 min at 3
and produced 39 times higher proteolytic activity than A. oryzae O-1. With protoplast fusion between A. oryzae O-1 and A. oryzee E-1 in the presence of polyethylenegylcol (PEG)-CaCl
, proteolytic activity was increased to 82 times compared to A. oryzae O-1, and the fusant was named A. oryzae PF. The activities of the cultures containing proteolytic enzymes produced by the strains were determined to be 0.23 U/
for A. oryzae O-1, 3.29 U/
for A. oryzae U-1, 8.91 U/
for A. oryzae E-1, and 19.0 U/
for A. oryzae PF.
Expression in Eschepichia coli of a Cloned Bacillus thuringiensis subsp. kurstaki HDI In-secticidal Protein Gene.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 497~506
The expression in Escherichia coli of a cloned insecticidal protein (ICP) gene from Bacillus thuringiensis var. kurstaki HD1 in pHLN1-80 (+) and pHLN2-80(-) plasmids was investigated through deletions in promoters, transcription start point, and termination region. Six recombinant plasmids were constructed in an attempt to analyze the overexpression of the ICP in relations to its gene structure. The amounts of ICP produced from the recombinants were measured by SDS-PAGE and confirmed by Western blot analysis. One clone was not overexpressed which having only -80 bp (contained BtI promoter) part of the ICP gene promoter (without Plac promoter), the right-oriented ICP gene and the termination region. Removal of 350 bp from upstream region of the Plac of the clone pHLN2-80 (-) resulted in overexpression of the ICP. One clone was not overexpressed in which the clone consisted of -72 bp part of the ICP promoter without the transcription start point and the transcriptional termination region, and having the right-oriented ICP gene sequence. One clone consisting of the inverted ICP gene sequence, the -72 bp ICP gene promoter, and without the termination region caused overexpression. One clone which consisted of the inverted ICP gene, the -72 bp ICP gene promoter and the termination sequence was overexpressed. These results indicated that the Plac promoter, transcription termination region, the inverted ICP gene insertion, and the -80 bp or -72 bp part of the ICP gene promoters were concerned in the overexpression of the ICP gene in the recombinant plasmid, and also the overexpression mechanism might result from the disruption of the transcription-suppressing regions in the promoter regions.
Purification and Characterization of Fibrinolytic Enzyme Excreted by Bacillus subtilis K-54 Isolated from Chung Guk Jang.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 507~514
The strain K-54, the best producer of fibrinolytic enzyme, was isolated from Korean traditional food Chung Guk Jang and identified as Bacillus subtilis. Fibrinolytic enzyme was purified and characterized, and its molecular weight was determined. The fibrinolytic enzyme activity was increased about 66.9 times via purification with recovery yield of 10.1%. The optimum pH and temperature of this enzyme were 11 and
. The enzyme was stable within a pH range 8-12 and unstable at 9
. The molecular weight was estimated to be 29,000 dalton in the form of monomer with no other subunit. The isoelectric point was calculated 8.67. N-terminal sequence was identified Ala-Gly-Ser-Val-Pro-Try-Gly-Ser. Km value of the enzyme for
-casein was calculated to be 0.31 (3.1 mg/
). The enzyme activity highly inhibited by PMSF at 1 nM.
Purification and Characterization of Chitinase from Antagonistic Bacteria Pseudomonas sp. 3098.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 515~522
Plant root rotting fungi, Fusarium solani are suppressed their growth by the chitinase which is produced from the antagonistic soil bacteria. The chitinase producable antagonistic bacterium Pseudomonas sp. 3098 was selected as a powerful biocontrol agent of F. solani from ginseng rhizosphere. The antagonistic Pseudomonas sp. 3098 was able to produce a large amount of extracellular chitinase which is key enzyme in the decomposition of fusarial hypal walls. The chitinase was purified from cultural filtrate of Pseudomonas sp. 3098 by the procedure of ammonium sulfate precipitation, anion exchange chromatography, gel filtration on Bio-Gel P-100, and 1st and 2nd hydroxyapatite chromatography. The molecular mass of the purified enzyme was ca. 45 kDa on SDS-FAGE. The optimal pH and temperature for the activity of purified chitinase were 5.0 and 45
, respectively. The enzyme was stable in pH range of 5.0 to 9.0 up to 5
The enzyme was significantly inhibited by metal compounds such as FeCl
, and was slightly inhibited by p-CMB, iodoacetic acid, urea, 2,4-DNP and EDTA. The enzyme had ability of digestion on colloidal chitin and chitin from shrimp shell, but could not digest chitosan and chitin from crab shell. Km value of the enzyme was 0.11% on colloidal chitin, and the maximum hydrolysis rate of the enzyme was 34% on colloidal chitin.
Purification and Characterization of Bacteriocin Produced by Enterococcus sp.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 523~528
We isolated microorganism secreting antimicrobial substance from tomato and identified as Enterococcus faecium. This substance was completely inactivated by pretense treatment and retained activity after catalase treatment. This result indicated that the antimicrobial activity of this substance was due to proteinaceous substance known as bacteriocin. The bacteriocin inhibited growth of Gram positive bacteria, such as Listeria monocytogenes, Leuconostoc mesenteroides, Lactobacillus plantarum, Streptococcus agalactiae, Streptococcus pyrogenes, and Gram negative bacteria, such as Pseudomonas aeruginosa. Purification of the bacteriocin was achieved by ethanol precipitation, ion exchange chromatography on CM Sepharose CL-6B, and gel filtration on Sephacryl S-100 HR. After these purification steps, the specific activity of the bacteriocin was increased 35.8 fold compared with culture broth. Purified bacteriocin was shown single band on SDS-PAGE and molecular weight was estimated 51 kDa. The residual activity of this bacteriocin was 3.3% at 10
for 60 min, and this bacteriocin was stable at pH 2~7.
Effect of Oxygen Transfer Rate and Dissolved Oxygen on the Production of PHBV by Azoto-bacter vinelandii UWD.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 529~536
In a 20 L fermentor experiments the level of dissolved oxygen (D.O.) strongly affected growth and PHBV production of Azotobacter vinelandii UWD. A higher D.O. (5%) increased specific cell growth rate two folds but PHBV production was 17 folds higher (62.3 wt%) at a lower D.O.(1%) level. D.O. level was not a good criterion to evaluate the effect of aeration on fermentation characteristics of A. vinelandii UWD. This strain maintained an equal D.O. (5%) by decreasing its oxygen consumption rate when oxygen transfer rate (OTR) was decreased by changing agitation speed at a fixed aeration rate. OTR rather than D.O. was a criterion to explain the effect of aeration on the cell growth and PHBV production. At 5% D.O. with a lower 0TR cell growth rate decreased but PHBV production (57.3 wt%) approached to that (62.3 wt%) of the lower (1%) D.O.
Cultivation Condition of Transformant Alcaligenes eutrophus Harboring Cloned phbC Gene for Production of P(3-hydroxybutyrate-3-hydroxyvalernte) Containing High Molar Fraction of 3-Hydroxyvalerate.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 537~544
The cultivation conditions of transformant Alcaligenes eutrophus AER5 harboring cloned phbC gene for mass production of poly (3-hydroxybutyrate-3-hydroxyvalerate)[P(3HB-3HV)] containing high molar fraction of 3-hydroxyvalerate (3-HV) were investigated. In two-stage batch cultivation, transformant accumulated P(3HB-3HV) containing 52.2 mol% of 3HV compared to 30 mol% of parent strain A. eutrophus H16. The increased 3-HV molar fraction was due to the amplified activity of PHB synthase participating in condensation of 3-HB and 3-HV. To increase efficiency of P(3HB-3HV) accumulation, fructose was added along with precursor compound valerate, and total cell mass and P(3HB-3HV) concentrations remarkably increased, but not 3-HV molar fraction. The effect of magnesium ion showed that P(3HB-3HV) concentration and 3-HV molar fraction were significantly increased upto 6.1 g/L and 71.3 mol% at 0.01 g/L of MgSO
, respectively. The efficiency of several pH adjuster, NaOH, NaOH and (NH
, and NH
OH, on total cell mass, p(3HB-3HV) concentration, and 3-HV molar fraction was also compared. To overcome the disadvantage of two-stage cultivation, one-stage intermittent fed-batch cultivation was attempted, such a way 10.0 g/L of fructose was supplied for cell growth at initial 36 hr and then 10.0 g/L of valerate and 5.0 g/L of fructose were applied to induce the accumulation of P(3HB-3HV), consequently, 10.4 g/L of P(3HB-3HV) with 38 mol% of 3-HV fraction could be obtained after 72 hr. These results can be used for elucidating cultivation strategy for mass production of P(3HB-3HV) containing high 3-HV molar fraction using transformant A. eutrophus AER5 harboring cloned phbC gene.
Isolation and Characterization of the Enterococcus sp. RKY1 for Biosynthesis of Succinic Acid.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 545~550
Succinic acid, valuable
-dicarboxylic acid as a renewable alternative feedstock, is currently produced commercially by the petrochemical process, but extensive efforts have been devoted to establish the biological process for mass production of succinic acid. In this study, the bioconversion of fumaric acid to succinic acid was investigated. We isolated an Enterococcus sp. RKY1 KCTC 8890P, facultative bacterium, capable of the bioconversion of fumaric acid to soccinic acid very rapidly and efficiently. At batch fermentation, the amount of succinic acid production increased with increase in initial fumaric acid from 40 to 100 g/L. With fumaric acid of 70 g/L, the average specific and volumetric production rate, molar yield were reached up to 0.64 g/g.h, 4.87 g/g.h, and 96.5%, respectively. Maximum concentration of succinic acid of 88.9 g/L was achieved with molar yield of 89% with fumaric acid of 100 g/L in less than 20 hours.
Biochemical Characteristics of Whole Soybean Cereals Fermented with Aspergillus Strains.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 551~557
Whole soybean cereal was fermented with four Aspergillus strains in pilot meju fermentation system. The pH range of the product was 7.40~7.98, the contents of reducing sugar and amino-nitrogen were 0.04~2.78%, 178~309 mg%, respectively and that of free fatty acid ranged 2.67~5.05%. The components of the amino acid, organic acid, free sugars and fatty acid showed distinctive patterns among four groups of fermented soybean cereals. Amylase activity and carbohydrate degradation rate of A. usami was higher than other strains. But protease and protein degradation rate, lipase and lipid degradation rate were similar in four strains. The odor concentrates of soybean cereals fermented with Aspergillus strains were different from Bacillus strains. Especially, pyrazine components, the main and common flavor chemicals in Bacillus strains, were not determined in this study and Aspergillus specific components were 9-methyl-acridine, dl-limonene and 2,3-butanediol. Soybean paste, made from A. oryzae fermented soybean cereal, showed excellent sensory evaluation.
A Nucleoside with Lipid Peroxidation Inhibitory Activity from Agrocybe cylindracea.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 558~561
In the screening for lipid peroxidation inhibitors from edible mushroom, Agrocybe cylindracea, a bioactive compound AG 8 was isolated. The AG 8 was purified from methanol extract of its fruit body by Diaion HP-20 column chromatography, ethyl acetate extraction, and silica gel column chromatography, consecutively. Based on various NMR studies including
H irradiation and HMBC experiments, the AG 8 was identified as MTA, 5'-deoxy-5'-methylthioadenosine. This compound inhibited lipid peroxidation with an
/ value of 3.2
. The MTA was isolated for the first time from basidiomycetes.
Characteristics of the Plasmid pCS100 Containing Nisin Resistant Gene from Lactococcus lactis subsp. lactis ATCC7962.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 562~565
Nisin-producing and nisin resistant L. lactis subsp. lactis ATCC7962 harbored six plasmids. To find a plasmid containing a nisin resistant gene, these plasmids were transformed into L lactis LM0230 of plasmid-free and nisin sensitive strain. After screening on nisin selection media containing nisin (150
), several nisin resistant transformants were obtained and the level of nisin resistance was very similar to that of wild type L lactis subsp. lactis ATCC7962. A 26.5 kb plasmid, named as pCS100, which confers resistance to nisin, was identified in transformants. The pCS100 was digested with EcoRI and Southern blot hybridization was done with nisI probe to localize the nisin resistant gene. A 4 kb EcoRI fragment showed a strong positive signal, and it was cloned into pBluescript for the potential selection marker.
Nucleotide Sequence of Pre Protein in Chloramphenicol Resistance Plasmid pKH7.
Microbiology and Biotechnology Letters, volume 26, issue 6, 1998, Pages 566~568
Partial nucleotide sequence (nt 1-1842) of chloramphenicol resistance plasmid pKH7 has been reported previously and residual nucleotide sequence (nt 1843-4118) of pKH7 was determined and then the complete nucleotide sequence of pKH7 was obtained. pKH7 consists of 4118 bp and has three ORFs. Besides Rep and CAT proteins described in previous paper, Pre protein which mediates site-specific recombination in Staphylococcus aureus was found to be on pKH7. R
, a site-specific recombination site of Pre protein, and palA, a specific lagging-strand conversion signal, was also found in pKH7. Amino acid sequence of Pre protein of pKH7 was compared with those of other antibiotic resistant Staphylococcus aureus plasmids.s.