• KSBB Journal
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• v.16 no.2
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• pp.188-199
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• 2001

A gene encoding the dextransucrase(dsCB) that synthesizes mostly $\alpha-(1\rightarrow6)$ linked dextran with low amount(10%) of $\alpha-(1\rightarrow3)$ branching was cloned and sequenced from Leuconostoc mesenteroides B-742CB. The 6.1 kbp DNA fragment carrying dsCB showed one open reading frame(ORF) composed of 4,536bp. The deduced amino acid sequence shows that it begins from the start codon(ATG) at position 698 of the cloned DNA fragment and extends to the termination condon(TAA) at position 5,223. The enzyme is consisted of 1,508 amino acids and has an calculated molecular mass of 168.6kDa. This calculated Mw was in good agreement with an activity band of 170kDa on non-denaturing SDS-PAGE. A recombinant E. coli DH5 $alpha$ harboring pDSCB produced extracellular dextransucrase in 2% sucrose medium, and synthesized both soluble and insoluble dextran. To compare the properties of enzyme with B-742CB dextransucrase, the acceptor reaction, hydrolysis of dextran and methylation were performed. The expressed enzyme showed the same properties as B-742CB dextransucrease, but its ability to synthesize $\alpha-(1\rightarrow3)$ branching was lower than that of B-742CB dextransucrase. In order to identify the critical amino acid residues known as conserved regions related to catalytic activity, Asp-492 was replaced with Asn. D492N resulted in a 1.6 fold decrease in specific activity.

• Journal of Microbiology and Biotechnology
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• v.14 no.3
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• pp.584-592
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• 2004

A thermostable $\beta$-glycosidase gene, tfi $\beta$-gly, was cloned from the genomic library of Thermus filiformis Wai33 A1. ifi $\beta$-gly consists of 1,296 bp nucleotide sequence and encodes a polypeptide of 431 amino acids. It shares a strong amino acid sequence similarity with the $\beta$-glycosidases from other Thermus spp. belonging to the glycosyl hydrolase family 1. In the present study, the enzyme was overexpressed in Escherichia coli BL21 (DE3) using the pET21b(+) vector system. The recombinant enzyme was purified to homogeneity by heat treatment and a $Ni^{2+}$-affinity chromatography. Polyacrylamide gel electrophoresis (PAGE) showed that the recombinant Tfi $\beta$-glycosidase was a monomeric form with molecular mass of 49 kDa. The temperature and pH range for optimal activity of the purified enzyme were 80- $90^{\circ}C$ and 5.0-6.0, respectively. Ninety-three percent of the enzyme activity was remained at $70^{\circ}C$ after 12 h, and its half-life at $80^{\circ}C$ was 6 h, indicating that Tfi $\beta$-glycosidase is highly thermostable. Based on its K_m$, or $K_{cat}K_m$, ratio, Tfi $\beta$-glycosidase appeared to have higher affinity for $\beta$-D-glucoside than for $\beta$-D-galactoside, however, $K_{cat} for \beta$-D-galactoside was much higher than that for $\beta$-D-glucoside. The activity for lactose hydrolysis was proportionally increased at $70^{\circ}C$ and pH 7.0 without substrate inhibition until reaching 250 mM lactose concentration. The specific activity of Tfi TEX>$\beta$-glycosidase on 138 mM lactose at $70{^\circ}C$ and pH 7.0 was 134.9 U/mg. Consequently, this newly cloned enzyme appears to have a valuable advantage of conducting biotechnological processes at elevated temperature during milk pasteurization in the production of low-lactose milk.

• Microbiology and Biotechnology Letters
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• v.30 no.1
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• pp.1-7
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• 2002

Phytase from Escherichia coli WC7 was purified from cell extracts and its molecular mass was estimated to be 45 kDa by SDS-PAGE. Its optimum temperature and pH for phytate hydrolysis was 6$0^{\circ}C$ and pH 5.0, respectively. The enzyme was stable up to 6$0^{\circ}C$ and over broad pH range (pH 2-12). The enzyme had higher affinity for sodium phytate than p-nitrophenylphosphate (pNPP). That is, the apparent Km value for sodium phytate and pNPP were $0.15\pm$0.02 mM and 2.82$\pm$0.05 mM, respectively. The gene encoding the phytase was cloned in E. coli XL1-Blue. Sequence analysis showed an open reading frame of 1241 Up encoding a signal peptide (22 aa) and a mature enzyme (410 aa). WC7 phytase was expressed up to 17.5 U/ml in the transformed E. coli XL1-Blue/pUEP, which was 23-fold higher than the activity from wild strain.

• Microbiology and Biotechnology Letters
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• v.31 no.4
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• pp.389-394
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• 2003

In order to screen microorganisms producing phopholipase D (PLD) had high transphosphatidylation activity, about 1,000 Actinomycetes strains were isolated from the 63 soil samples, collected over 6 local area in Korea. When the hydrolytic activity in the supernatant was determined, 131 strains produced PLD more than 0.3 U/ml. Among 131 culture broths tested, 23 ones had transphosphatidylation activity higher than 20% and finally one strain (Actinomycetes KF 923), which had highest hydrolytic and transphophadylation activity, was selected. Actinomycetes KF923 showed the highest hydrolytic activity (13 U/ml) and phosphatidylation activity (95%) after 48 h fermentation using the P medium (yeast extract 1%, peptone 1%, glucose 1.5%, glycerol 1%, $CaCo_3$ 0.4%, pH 7.2). PLD was purified from the culture broth of Actinomycetes KF923 and the specific activity of purified PLD was 567 U/mg. The molecular weight of PLD was about 55 kD and the optimum pH and temperature were 6.0 and $60^{\circ}C$, respectively. The stability of PLD toward pH and temperature were high around pH 8.0 and below $40^{\circ}C$. Special metal ions were not necessary to the PLD activity.

• Journal of Life Science
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• v.15 no.4 s.71
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• pp.657-663
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• 2005

Collagenases are generally defined as enzymes that are capable of degrading the polypeptide backbone of native collagen under conditions that do not denature the protein. An extracellular collagenase-producing bacterial strain was isolated from kimchi and identified to be Bacillus subtilis JS-17 through morphological, cultural, biochemical characteristics and 16S rDNA sequence analysis. Optimum culture condition of Bacillus subtilis JS-17 for the production of collagenase was $1.5\%$ fructose, $1\%$ yeast extract, $0.5\%\;K_2HPO_4,\;0.4\%\;KH_2PO_4,\;0.01\%\;MgSO_4\cdot7H_2O,\;0.01\%\; MnSO_4\cdot4H_2O,\;,0.1\%$ citrate and $0.1\%\;CaCl_2$. The production of collagenase was optimal at $30^{\circ}C$ for 72 hr. A collagenase was isolated from the culture filtrate of Bacillus subtilis JS-17. The enzyme was purified using Amberlite IRA-900 column chromatography, Sephacryl S-300 HR column chromatography and DEAE-Sephadex A-50 column chromatography The purified collagenase has an specific activity 192.1 units/mg. The molecular weight of the purified enzyme was estimated to be 28 kDa by SDS-PACE. The purified collagenase has $100\%$ activity up to $55^{\circ}C$.

• The Transactions of the Korean Institute of Electrical Engineers C
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• v.48 no.5
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• pp.285-295
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• 1999

Electroluminescent(EL) devices based on organic materials have been of great interest due to their possible applications for large-area flat-panel displays. They are attractive because of their capability of multi-color emission, and low operation voltage. An approach to realize such device characteristics is to use active layers of lanthanide complexes with their inherent extremely sharp emission bands in stead of commonly known organic dyes. In general, organic molecular compounds show emission due to their $\pi$-$\pi*$ transitions resulting in luminescence bandwidths of about 80 to 100nm. Spin statistic estimations lead to an internal quantum efficiency of dye-based EL devices limited to 25%. On the contrary, the fluorescence of lanthanide complexes is based on an intramolecular energy transfer from the triplet of the organic ligand to the 4f energy states of the ion. Therefore, theoretical internal quantum efficiency is principally not limited. In this study, Powders of TPD, $Eu(TTA)_3(phen) and AlQ_3$ in a boat were subsequently heated to their sublimation temperatures to obtain the growth rates of 0.2~0.3nm/s. Organic electrolumnescent devices(OELD) with a structure of $glass substrate/ITO/Eu(TTA)_3(phen)/AI, glass substrate/ITO/TPD/Eu(TTA)_3(phen)/AI and glass substrate/ITO/TPD/Eu(TTA)_3(phen)/AIQ_3AI$ structures were fabricated by vacuum evaporation method, where aromatic diamine(TPD) was used as a hole transporting material, $Eu(TTA)_3(phen)$ as an emitting material, and Tris(8-hydroxyquinoline)Aluminum$(AlQ_3)$ as an electron transporting layer. Electroluminescent(EL) and current density-voltage(J-V) characteristics of these OELDs with various thickness of $Eu(TTA)_3(phen)$ layer were investigated. The triple-layer structure devices show the red EL spectrum at the wavelength of 613nm, which is almost the same as the photoluminescent(PL) spectrum of $Eu(TTA)_3(phen)$.It was found from the J-V characteristics of these devices that the current density is not dependent on the applied field, but on the electric field.

• Microbiology and Biotechnology Letters
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• v.48 no.2
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• pp.215-222
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• 2020

The marine microorganism PX-1, which can hydrolyze xylan, was isolated from coastal sea water of Jeju Island, Korea. Based on the 16S rRNA gene sequence and chemotaxonomy analysis, PX-1 was identified as a species of the genus Aestuariibacter and named Aestuariibacter sp PX-1. From the culture broth of PX-1, an extracellular xylanase was purified to homogeneity through ammonium sulfate precipitation and subsequent adsorption chromatography using insoluble xylan. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography estimated the molecular weight of the purified putative xylanase (XylA) as approximately 64 kDa. XylA showed xylanase activity toward beechwood xylan, with a maximum enzymatic activity at pH 6.0 and 45℃. Through thin-layer chromatographic analysis of the xylan hydrolysate produced by XylA, it was confirmed that XylA is an endo-type xylanase that decomposes xylan into xylose and xyloligosaccharides of various lengths. The K_m and V_max values of XylA for beechwood xylan were 27.78 mM and 78.13 μM/min, respectively.

• Journal of Korean Society of Environmental Engineers
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• v.33 no.11
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• pp.847-854
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• 2011

To deal with issues from global climate changes, renewable bioenergy has become important. Algae have been regarded as a good resource for biorefinery and bioenergy, and also have potential capability to remove nutrient and non-decompositional pollutants for wastewater advanced treatment. Although algal-bacterial ecological interaction would be a crucially important factor in using algae for wastewater advanced treatment and resource recovery from wastewater, very little is known about ecological interaction between algae and bacteria in a real wastewater environment. In this study, under a real municipal wastewater condition, we characterized wastewater pollutant treatability and bacterial communities in response to growth of Ankistrodesmus gracilis SAG278-2, which can grow in wastewater and has a high lipid contents. The growth of algal population using the wastewater was inhibited by increase in wastewater bacteria while bacterial survival and cellular decay rate were not influenced by the algal growth. Removals of recalcitrant organic matters and total nitrogen were improved in the presence of algal growth. According to T-RFLP and statistical analysis, algal growth affected time-course changes in bacterial community structures. The following 16S rRNA gene amplicon, cloning results showed that the algal growth changes in bacterial community structure, and that bacterial populations belonging to Sediminibacterium, Sphingobacterium, Mucilaginibacter genera were identified as cooperative with the algal growth in the wastewater.

• Analytical Science and Technology
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• v.11 no.1
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• pp.73-78
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• 1998

The reverse detection heteronuclear multiple quantum coherence, HMQC study of metcyano complex of horse myoglobin(MbCN) has provided the complete assignment of hyperfine shifted resonances of heme carbons attached with proton(s). The application of HMQC experiment to the paramagnetic low-spin MbCN gives clear $^1H$ and $^{13}C$ coherences for the paramagnetic amino acid residues as well as heme side chains, and can be extended to the low-spin paramagnetic hemoprotein derivative for the assignment of natural abundance $^{13}C$ resonances. This assignment strategy can avoid possible ambiguities that may result from the sole utilization of $^1H$ nuclear Overhauser effect for the assignment of heme $^1H$ signals resonating in the diamagnetic region. The resulting 2,4-vinyl ${\alpha}$-carbons and 7-propionate ${\beta}$-carbon follow anomalous anti-Curie behavior, and are indicative of incoplanarity with heme plane. Magnetic/electronic asymmetry of heme induced by proximal histidine(His) makes spread that the hyperfine shifted heme carbon resonances over the range of 250 ppm at $25^{\circ}C$. These heme carbon resonances would be the much more sensitive probe than those of proton resonances in analyzing the nature of heme electronic structure of myoglobin.

• The Korean Journal of Mycology
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• v.42 no.4
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• pp.262-268
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• 2014

A fungal isolate DUCC5000 from a garden plant Pachysandra terminalis was identified as Paraconiothyrium brasiliense based on the results of morphological and molecular studies. The fungus formed brown to black conidiomata of (0.2-0.7)-2(-3.5) mm singly or as a group on PDA. Conidia measured $2-5{\times}1.8-3{\mu}m$ in size, hyaline, ellipsoid to short-cylindrical, and rounded at both ends. The internal transcribed spacer (ITS) DNA of the isolate shared 100% nucleotide sequence homology with those of known P. brasiliense isolates. Phylogenetic tree inferred from the ITS sequence analysis showed that the DUCC5000 isolate formed a clade with known isolates of P. brasiliense. The fungal mycelia grew better on oatmeal agar than on MEA and PDA. On PDA media under various pH conditions, fungal mycelial growth was observed at pH 9. Colony morphology of the fungus tended to alter depending on the kinds of nutrient media and pH condition. On chromagenic media, the fungus demonstrated its ability to produce extracellular enzymes including amyalse, avicelase, ${\beta}$-glucosidase, protease, and xylanase. However, in pathogenicity testing, no disease symptoms were observed on the leaves of P. terminalis. This strain is the first report on P. terminalis in Korea.