• Maxillofacial Plastic and Reconstructive Surgery
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• v.19 no.3
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• pp.265-286
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• 1997

Bone morphogenetic proteins(BMPs) are a group of transforming growth factor beta(TGF-${\beta}$)-related factors and multifunctional proteins, especially the only known biologic factors capable of inducing endochondral bone formation at an extraskeletal site. This study was performed to investigate the effect of the partially purified porcine BMP(pBMP) at an ectopic site. PBMP was partially purified from porcine bone matrix and its activity was monitored by an in vivo bioassay. The purification method utilized extraction of the bone-inducing activity with 4M guanidine, followed by chromatography on heparin-Sepharose. Active fractions were assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. And the fractions were reconstituted with inactive insoluble collagenous bone matrix from rats, acid soluble type I collagen from rat tail and chondroitin-6-sulfate sodium salt and implanted into the pectroralis muscle pouches of Sprague-Dawley rats. And the carrier complex was implanted on the opposite side as control. The rats were sacrificed at the day of 1st, 3rd, 5th, 7th, 11th, 14th and 21st after implantation and examined histologically, radiologically and biochemically. And alkaline phosphatase activity and calcium content were used as indices of bone formation. The results were as follows ; 1. Active fractions were localized in a zone between 31 and 40 KDa on SDS-PAGE. 2. The implanted 3.0mg of the partially purified pBMP induced cartilage and bone in the muscle tissue of rats through an endochondral ossification process. 3. Inactive insoluble bone matrix, type I collagen and chondroitin-6-sulfate have functioned as carriers for pBMP, but revealed some foreign body reactions. 4. Soft X-ray didn't reveal significant change between the experimental and the control group. 5. The alkaline phosphatase activities in the experimental group of 5th, 7th, 11th, 14th and 21st were increased significantly compared with control (p<0.01) with the peak in the group of 11th day. 6. With time, the calcium content of the experimental group increased. And the calcium contents in the experimental group of 11th, 14th and 21st were increased significantly compared with control (p<0.01).

• The Korean Journal of Food And Nutrition
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• v.15 no.2
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• pp.104-111
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• 2002

The peptidyl prolyl sis-trans isomerase (PPIase, EC 5.2.1.8) from bacillus stearothermophilus was extracted from the cells treated with by lysozyme. PPIase was purified from the cell extracts by heat treatment, ammonium sulfate precipitation, ion exchange chromatography and finally gel filtration, sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). The molecular weight of the purified PPIase was estimated as 18kDa by SDS-PAGE. The 39 amino acid residues from the N-terminus were determined by the protein sequencer. The enzyme showed the optimum pH at 8.0 and was stable at the range of pH 7.0∼8.0. The enzyme was considerably stable after heat treatment at 60$\^{C}$ for 30minutes, and the enzyme was quite stable up to 65$\^{C}$. The presence of the PPIase in the refolding solution accelerated the isomerization rate of the assay peptide. PPIase gene of Bacillus stearothermophilus was screened from a genomic library by plaque hybridization using the A-l primer as a probe. A PPIase positive plaque contained a 3.0kb insert of the chromosomal DNA. A 3.0kb fragment was subcloned into pUC18, resulting pPI-40. A DNA fragment encoding the N-terminal portion of the PPIase in pPI-40 was amplified by polymerase chain reaction(PCR) method using the A-1 and B-2 primers. The amplified fragment was cloned into the Sma I site of pUC18 and recombinant plasmid was designated as pSN-18. The nucleotide sequence of 167bp fragment was determined. The deduced amino acid sequence of PPIase was completely matched with the determined N-terminal amino acid sequence of PPIase B. stearothermophilus.

• Microbiology and Biotechnology Letters
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• v.48 no.4
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• pp.463-470
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• 2020

The JS18 strain was isolated from an old tree forest and produced extracellular enzymes that decolorize synthetic melanin. Phylogenetic analysis, based on the internal transcribed spacer (ITS) sequence, indicate that JS18 belongs to the Trametes velutina species. JS18 demonstrated laccase activity but no manganese peroxidase or lignin peroxidase activity. Batch culture indicated that the melanin decolorization activity of JS18 strain originated from the laccase. Syringic acid and CuSO4 induced maximum laccase production, yielding 98 U/ml laccase activity after cultivation for 7 days at 25℃. T. velutina secretes an extracellular laccase in GYP medium, and this enzyme was purified using (NH4)2SO4 precipitation, Hi-trap Q Sepharose columns and gel filtration. The molecular weight of the purified enzyme was estimated to be 67 kDa using sodium dodecyl sulfate polyacrylamide gel electrophoresis. This enzyme produced 80% of its melanin decolorization activity within the first 24 h of evaluation in the presence of 1-hydroxybenzotriazole (HBT), while only about 4% of the melanin was decolorized in the absence of the mediator. The greatest decolorization was observed at 1.5 mM/l HBT, which decolorized 81% of the melanin within the first 24 h. The optimum pH and temperature for this decolorization were found to be 5.0 and 37℃, respectively. Our results suggest the possibility of applying HBT induced T. velutina JS18 laccase-catalyzed melanin decolorization.

• Journal of Life Science
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• v.13 no.1
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• pp.90-98
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• 2003

An allium rhizobacterium Serratia plymuthica AL-1 was previously selected as a biocontrol agent of allium white rot. The chitinase from S. plymuthica AL-1 produced in medium containing colloidal chitin was purified by ammonium sulfate precipitation (40~70%), affinity adsorption, column chromatography on DEAE-sephadex A-50 and sephadex C-200 gel filtration. The enzyme was purified 10.8-fold with a yield of 7.3% from the starting culture broth. The purified chtinase gave a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis, it's molecular weight was estimated to be 55 kDa. The optimum pH and temperature of the purified enzyme were pH 5.5 and $55^{\circ}C$, respectively and it is stable up to $50^{\circ}C$ and maintains around 90% of its activity for 60min. The enzyme were activated by $Ca^{2+}$, $Mn^{2+}$ and $Mg^{2+}$ and inhibited by $Cu^{2+}$, SDS, $\rho$-CMB, MIA, respectively. The purified chitinase showed broad spectrum of antifungal activities against plant pathogenic fungi Sclerotium cepivoruin, Alternana alternnta, Colletotrichum glceosporioidrs, Phoma sp., Sclerotinia sclerotiorum, Stemphylium solani, Fusarium oxysporium f. sp. niveum but rarely inhibited Phytophthora capsici and Pythium ultimum.. The purified chitinase from S. plymuthica AL-1 caused swelling, lysis, deceleration and degradation of the hyphal tips of S. sczerotiorum causing allium white rot. It suggest that S. prymuthica AL-1 chitinase play an important part in the bifunctional chitinase / lysozyme activity.

• Microbiology and Biotechnology Letters
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• v.42 no.4
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• pp.393-401
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• 2014

A bacterium producing antimicrobial substance was isolated from cheonggukjang. The bacterium was identified as a strain of Bacillus subtilis by 16S rDNA sequencing and designated as Bacillus subtilis HH28. The antimicrobial substance produced from Bacillus subtilis HH28 was purified by 0-80% ammonium sulfate precipitation, DEAE-sepharose FF column chromatography, and Sephacryl S-200 HR gel chromatography. The molecular weight of the purified antimicrobial substance was estimated to be approximately 3,500 Da using Tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis and direct detection analysis. Antimicrobial substance from B. subtilis HH28 not only inhibited B. cereus, but also Listeria monocytogenes and Vibrio parahaemolyticus. The purified antimicrobial substance was stable at $40-80^{\circ}C$, and between pH 2 and 8. Antimicrobial activity of the purified substance was completely destroyed by treatment of protease, proteinase K, and pronase E, indicating that it is proteinaceous.

• Food Science of Animal Resources
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• v.32 no.3
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• pp.323-329
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• 2012

In order to investigate the rheological properties of myofibrillar protein (MP) mixed with tapioca starch (TS; 0, 1, and 2%) at various salt concentrations (0.1, 0.3, and 0.45 M), viscosity, gel strength, differential scanning calorimeter (DSC) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) were measured. Viscosity of MP increased with increasing salt concentrations (p<0.05), but not with the addition of TS. The addition of TS improved gel strength and cooking yield at all salt concentrations (p<0.05). DSC results demonstrated that the starting peak of TS gelation was observed at $55^{\circ}C$, however, no differences in peak were observed with various salt and TS levels (p>0.05). SDS-PAGE profile also showed no differences in protein bands for pork myofibrillar protein with various salt and TS levels. Based on the model study, pork model sausages with various levels of tapioca (0, 1, and 2%) and TG (1%) were manufactured. The pork model sausages with 2% TS increased pH and water holding capacity (p<0.05), while those with TGase (1%) increased most textural properties, regardless of the addition of TS. Thus, the combination of 1% TG with 2% TS improved the gel strength and water holding capacity in the meat products.

• Journal of Life Science
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• v.21 no.11
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• pp.1565-1572
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• 2011

Eye-specific lactate dehydrogenase (EC 1.1.1.27, LDH) $C_4$ isozyme in the eyes of greenlings (Hexagrammos otakii) was successfully purified by affinity chromatography and continuous-elution electrophoresis. The molecular weight of the purified eye-specific LDH $C_4$ isozyme was 154.8 kDa, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Optimal pH for enzymatic reaction of the eye-specific LDH $C_4$ isozyme was pH 8.5. $K^{PYR}_m$ value of the purified eye-specific LDH $C_4$ isozyme was $1.88{\times}10^{-5}$ M using pyruvate as a substrate. These results indicate that we must consider pH when measuring eye-specific LDH $C_4$ isozyme activity. The eye-specific LDH $C_4$ isozyme had a higher binding affinity for the substrate as a pyruvate than LDH A4 isozyme. Antibodies produced against the purified eye-specific LDH $C_4$ isozyme may be used in the diagnosis of several human diseases and in comparative physiological studies of fishes.

• Asian-Australasian Journal of Animal Sciences
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• v.23 no.3
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• pp.347-354
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• 2010

This study aimed to evaluate effects of 800 W microwave irradiation for 2, 4 and 6 min on chemical composition, antinutritional factors, ruminal dry matter (DM) and crude protein (CP) degradability, and in vitro CP digestibility of canola seed (CS). Nylon bags of untreated or irradiated CS were suspended in the rumen of three bulls from 0 to 48 h. Protein subfractions of untreated and microwave irradiated CS before and after incubation in the rumen were monitored by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Microwave irradiation had no effect on chemical composition of CS (p>0.05). There was a linear decrease (p<0.001) in the phytic acid and glucosinolate contents of CS as irradiation time increased. Microwave irradiation for 2, 4 and 6 min decreased the phytic acid content of CS by 8.2, 27.6 and 48.6%, respectively. The total glucosinolate contents of CS microwave irradiated for 2, 4 and 6 min decreased by 41.5, 54.7 and 59.0% respectively, compared to untreated samples. The washout fractions of DM and CP and degradation rate of the b fraction of CP decreased linearly (p<0.001) as irradiation time increased. Microwave irradiation for 2, 4 and 6 min decreased effective degradability (ED) of CP at a ruminal outflow rate of 0.05 $h^{-1}$ by 4.7, 12.3 and 21.0%, respectively. Microwave irradiation increased linearly (p<0.001) in vitro CP digestibility of ruminally undegraded CS collected after 16 h incubation. Electrophoresis results showed that napin subunits of untreated CS disappeared completely within the zero incubation period, whereas cruciferin subunits were degraded in the middle of the incubation period (16 h incubation period). In 4 and 6 min microwave irradiated CS, napin subunits were degraded after 4 and 16 h incubation periods, respectively, and cruciferin subunits were not degraded untile 24 h of incubation. In conclusion, it seems that microwave irradiation not only protected CP of CS from ruminal degradation, but also increased in vitro digestibility of CP. Moreover, microwave irradiation was effective in reducing glucosinolate and phytic acid contents of CS.

• Korean Journal Plant Pathology
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• v.1 no.1
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• pp.3-11
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• 1985

Three mild mutant strains of tobacco mosaic virus (TMV) were isolated from Nicotiana tabacum var. Samsun incubated at $38^{\circ}C$ for 10 days after inoculation with a wild type of TMV-OM strain. They were designated into Tg 5272, Tw 227 and Tw 333. All mild strains could be distinguished from TMV-OM by their reactions on different indicator plants. The mild strains induced the mild mottling without distinct symptoms, whereas the wild strain produced severe mosaic, rugose and stunting on tobacco and red pepper plants. Tw 227 and Tw 333 produced smaller necrotic spots than those of Tg 5272 and TMV-OM on N. glutinosa and Datura stramonium. The former two strains also produced ring spots and mosaic on Gomphrena globosa compared with necrotic spots by the latter strains. Three mild strains were serologically identical to TMV-OM. Their physical properties were thermal inactivation point $80-85^{\circ}C$, dilution end point between $10^{-4}\;and\;10^{-6}$, and longevity in vitro 7days or longer. Ultraviolet absorption spectra of purified preparations of the mild strains and TMV-OM were identical, with a minimum at 247nm, a maximum at 260nm, and a slight shoulder at 290nm. Electrophoresis of the strains in polyacrylamide-agarose gel showed that all the strains formed one major band and two minor bands, except for one minor band of Tw 333. However, when sodium dodecyl sulfate was added to the purified viruses before electrophoresis, each strain formed only one major band.

• Journal of Veterinary Clinics
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• v.25 no.3
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• pp.152-158
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• 2008

We investigated the changes of protein in chronic MI which was occurred with long-term ischemia, without reperfusion. Sprague Dawley (SD) rats were divided into the sham group and the experimental groups (MI groups). The sham group was treated only thoracotomy without ligation for left main descending artery (LMDA) of left coronary artery (LCA), and the experimental groups (MI7d, ligation of LMDA for 7 days and MI30d, ligation of LMDA for 30 days) were conducted an artificial chronic MI. The change of proteins according to passage of times was compared and analyzed on first and second dimension (1 and 2D) sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. Among total 46 spots expressed differentially in the sham group versus MI7d and MI30d groups on 2D gel, we selected proteins that the volume of spot was increased in the MI7d and MI30d groups compared with the sham group. After that, the proteins were identified through liquid chromatography/tandem mass spectrometry (LC-MS/MS) analysis. In result, we could obtain many proteins as follows; albumin, glucose regulated protein 58 KDa, similar to tripartite motif protein 50, ubiquinol-cytochrome c reductase core protein II, sarcomeric mitochondrial creatine kinase, ATP synthetase alpha chain (mitochondrial precursor) and creatine kinase. In conclusion, we suggest many changed proteins shown at chronic ischemia after artificial MI and consider that these proteins play an important role in the function of heart after MI.