• Journal of Life Science
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• v.27 no.3
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• pp.295-300
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• 2017

This study examined the association between genotypes of the isocitrate dehydrogenase 3, beta subunit (IDH3B) gene and economic traits in an $F_2$ population of Duroc and Jeju (South Korea) native black pigs (JBPs). The genotypes was determined the presence/absence of a 304-bp insertion/deletion fragment in the promoter region of the IDH3B gene for JBP, Duroc, and their $F_1$ and $F_2$ progeny. Three genotypes (AA, AB and BB) were found in the $F_1$ and $F_2$ populations, but there was no AA genotype found in JBP and no BB in Duroc. Association analysis results showed the significant differences with carcass weights (CW), backfat thicknesses (BFT) and eye muscle area (EMA) (p<0.05), but not with growth traits including body weights and average daily gains at different stages, reproductive traits including teat numbers, and crude fat contents (CFAT) measured in longissimus dorsi (p>0.05). The $F_2$ pigs possessing the IDH3B BB homozygote had heavier CW ($72.92{\pm}11.133kg$), thicker BFT ($25.75{\pm}6.06mm$), and larger EMA ($23.82{\pm}4.825cm^2$) than those from the other genotypes (p<0.05). These results were estimated that there are biological roles related with IDH3B genotypes resulting development of EMA, BFT, and CW but not with intramuscular fat deposition during late period of pig production. Our findings suggest that the 304-bp insertion allele of porcine IDH3B may be a genetic marker for marker assistant selection for improving meat productivity of the Jeju Black pig and Duroc-related molecular breeding systems.

• Journal of Microbiology
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• v.45 no.2
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• pp.158-167
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• 2007

In this study, we have cloned a novel cDNA encoding for a papain-family cysteine protease from the Uni-ZAP XR cDNA library of the polychaete, Periserrula leucophryna. This gene was expressed in Escherichia coli using the T7 promoter system, and the protease was characterized after partial purification. First, the partial DNA fragment (498 bp) was amplified from the total RNA via RT-PCR using degenerated primers derived from the conserved region of cysteine protease. The full-length cDNA of cysteine protease (PLCP) was prepared via the screening of the Uni-ZAP XR cDNA library using the $^{32}P-labeled$ partial DNA fragment. As a result, the PLCP gene was determined to consist of a 2591 bp nucleotide sequence (CDS: 173-1024 bp) which encodes for a 283-amino acid polypeptide, which is itself composed of an 59-residue signal sequence, a 6-residue propeptide, a 218-residue mature protein, and a long 3'-noncoding region encompassing 1564 bp. The predicted molecular weights of the preproprotein and the mature protein were calculated as 31.8 kDa and 25 kDa, respectively. The results of sequence analysis and alignment revealed a significant degree of sequence similarity with other eukaryotic cysteine proteases, including the conserved catalytic triad of the $Cys^{90},\;His^{226},\;and\;Asn^{250}$ residues which characterize the C1 family of papain-like cysteine protease. The nucleotide and amino acid sequences of the novel gene were deposited into the GenBank database under the accession numbers, AY390282 and AAR27011, respectively. The results of Northern blot analysis revealed the 2.5 kb size of the transcript and ubiquitous expression throughout the entirety of the body, head, gut, and skin, which suggested that the PLCP may be grouped within the cathepsin F-like proteases. The region encoding for the mature form of the protease was then subcloned into the pT7-7 expression vector following PCR amplification using the designed primers, including the initiation and termination codons. The recombinant cysteine proteases were generated in a range of 6.3 % to 12.5 % of the total cell proteins in the E. coli BL21(DE3) strain for 8 transformants. The results of SDS-PAGE and Western blot analysis indicated that a cysteine protease of approximately 25 kDa (mature form) was generated. The optimal pH and temperature of the enzyme were determined to be approximately 9.5 and $35^{\circ}C$, respectively, thereby indicating that the cysteine protease is a member of the alkaline protease group. The evaluation of substrate specificity indicated that the purified protease was more active towards Arg-X or Lys-X and did not efficiently cleave the substrates with non-polar amino acids at the P1 site. The PLCP evidenced fibrinolytic activity on the plasminogen-free fibrin plate test.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.11
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• pp.1575-1581
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• 2005

This study was conducted to determine the characteristics of IGFBPs in plasma of steers, and to profile the relationship between growth and plasma IGF-1 and IGFBPs with aging in Holstein steers. Four blots of IGFBP at molecular weights of 38-43, 34, 29-32 and 24 kDa bands were detected by western ligand blot assay using $^{125}I-IGF-1$. On the basis of immunoblotting with anti-bovine IGFBP-2 and -3 antiserums, we observed the band for IGFBP-2 at approximately 34 kDa, and the IGFBP-3 band was detected at 38-43 kDa and 34 kDa in adult steers and calves. The IGFBP-3 antiserum used on the blots exhibited significant cross-reactivity with 34 kDa IGFBP-2. Furthermore, the 38-43 kDa IGFBP-3 bands were reduced to a 36 kDa band after deglycosylation, whereas the 34 kDa IGFBP-2 was intact. The plasma IGF-1, IGFBP-3 and other IGFBPs showed stability throughout a whole day. The change in live weight was found to be positively correlated to the plasma IGF-1 concentration (r = 0.6801, n = 64, p<0.05) and plasma IGFBP-3 (r = 0.6321, n = 64, p<0.05), while inversely correlated to plasma IGFBP-2 (r = -0.2919, n = 64, p<0.05). Furthermore, plasma IGF-1 was positively correlated to plasma IGFBP-3 (r = 0.6191, p<0.001), but was not correlated to plasma IGFBP-2. The portion of IGFBP-2 for total IGFBPs in calves was higher than in adult steers (p<0.05) and was decreased with growth, whereas that of IGFBP-3 was increased with increased live weight (p<0.05). The ratio IGFBP-3 for IGFBP-2 (BP-3/BP-2) was increased with growing of liveweight. Therefore, the changes in plasma IGF-1 level with increased liveweight may be related to the changes in plasma IGFBP-3 level and IGFBP-2 may give an important role in anabolic action of IGF-1 with the growth of body during calfhood in Holstein steers.

• Polymer(Korea)
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• v.36 no.6
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• pp.822-830
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• 2012

We used dipodal type bis[3-(trimethoxysilyl)propyl]amine (BTMA) silane coupling agent to modify silica nanoparticles to introduce secondary amino groups on the silica surface. These N-H groups were reacted with three different molecular weights (M.W. = 258, 575, and 700) of poly(ethylene glycol) diacrylates to introduce different attached layer thicknesses on the silica surface by Michael addition reaction. After Michael addition reaction, we used several analytical techniques such as fourier transform infrared spectroscopy (FTIR), elemental analysis (EA) and solid state $^{13}C$ cross-polarization magic angle spinning (CP/MAS) nuclear magnetic resonance spectroscopy to characterize introduced structures. We found almost complete Michael addition reaction of both two acrylate groups of PDGDA with N-H groups of BTMA modified silica to form ${\beta}$-amino acid esters. Between equimolar ratio of pure BTMA and pure PEGDA reaction, only one acrylate group of two acrylate groups of PEGDA reacted with N-H groups of pure BTMA to form ${\beta}$-amino acid ester and the other remaining acrylate group can be used to form a polymer later.

• Microbiology and Biotechnology Letters
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• v.44 no.2
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• pp.124-129
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• 2016

The extraction and purification of the anti-hyperglycemic α-glucosidase inhibitor from an edible mushroom, Pleurotus cornucopiae, were investigated. The inhibitor was maximally extracted when the P. cornucopiae fruiting body was treated with distilled water at 30℃ for 12 h. Purification was achieved using Sephadex G-100 and G-50 filtration chromatography, pepsin hydrolysis, and reverse-phase HPLC. The compound’s solid yield and inhibitory activity were 12.2% and 9.10 mg/ml of IC₅₀, respectively. The purified inhibitor contained two hexapeptides with Thr-Ile-Ala-Phe-Ile-Asp (A) and Tyr-Tyr-Ala-Ile-Gly-Asp (B) sequences and molecular weights of 678.79 Da (A) and 643.7 Da (B). The purified inhibitor showed a mixed inhibition pattern to α-glucosidase and a dose-dependent anti-hyperglycemic effect in a streptozotocininduced diabetic Sprague-Dawley rat model, exhibited by decreased blood glucose levels at doses of 50 and 300 mg/kg.

• Microbiology and Biotechnology Letters
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• v.26 no.6
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• pp.483-489
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• 1998

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.

• Journal of the Korean Society of Food Science and Nutrition
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• v.31 no.1
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• pp.1-6
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• 2002

Synthesis of new active protein was investigated by heat-treatment and fermentation of kidney beans with Bacillus subtilis ATCC 51189. The amount of water-soluble protein in raw kidney bean (raw protein, RP) was greatly reduced by heating (heated protein, HP) and several new amino acids were synthesized by fermentation. The molecular weights of proteins determined by SDS-PAGE were 118 kDa for RP and a new band of 18.0 kDa For protein (fermented protein, FP) in kidney beans heated and fermented with B. subtilis ATCC 51189. Hemagglutinating activities of RP, HP and FP were 128 HU, 4 HU and 32 HU respectively. Both of RP and FP showed anticancer activity against stomach cancer cell line (SNU-1) at 50 $\mu\textrm{g}$g/mL and lymphocyte stimulating activity at 1 $\mu\textrm{g}$/mL, and stimulated PBMC to secrete IFN-${\gamma}$ and IL-12. However, HP did not show any kinds of activities. Taken together, these results suggested that lectin in kidney beans was destroyed by heating, hut new active lectin-like Protein was derived by fermentation with B. subtilis ATCC 51189.

• KSBB Journal
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• v.19 no.4
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• pp.274-283
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• 2004

Six lumbrokinase (LK) fractions from Lumbricus rubellus lysates were purified by a series of column chromatographies. The molecular weights of the six LK fractions appeared to range from 24.6 to 33.1 kDa. In the experimental model of rat venous thrombosis, the thrombus weight and PAI activity decreased significantly when the LK was administered orally. However, the activities of APTT, PT and plasmin showed a significant increase. The aggregation of rat platelets pretreated with various LK doses was inhibited by thrombin, and the MDA generation decreased. The rat thoracic aorta and mesentric arteries contracted with phenylephrine relaxed due to the treatment of the LK fractions. These results suggest that the fibrinolytic effects of LK were mediated not only by proteolytic activity, but also by the inhibition of platelet agregation and the relaxation of blood vessels. It is concluded that the LK may be useful as a hemolytic agent for treatment of fibrin clot.

• KSBB Journal
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• v.14 no.5
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• pp.578-584
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• 1999

The lectin was purified through 0.15 M NaCl extraction, ammonium sulfate precipitation, sepharose 4B affinity chromatography and gel filtration using sephadex G-150 from the leaves of Visum album collected in Mt. Duk Yu. The final gel filtration step resulted in 11.64 folds purification with 0.14% of recovery yield. We also performed biochemical characterization of the purified Visum album lectin. HPLC analysis of lectin purified by gel filtration revealed a singel peak. The analysis of the purified lectin by SDS-PAGE showed a tetramer composed of two identical subunits with molecular weights of 32 and 30 kDa. The lectin was a glycoprotein containing 14.4% carbohydrate, which consist of glucose, fructose, arabinose and xylose, and the amino acids such as phenylalanine, lysine and tyrosine. The purified lectin agglutinated human red blood cell types with similar potency, but when tested against red blood cells from mouse, bovine, rabbit, chicken and porcine, significant difference in potency were observed. Hemaggluting activity was inhibited by D-galactose, D-mannose, D-lactose and D-raffinose, but not by D-glucose, D-glucosamine, D-mannosamine, L-fructose, D-xylose, D-arabinose, D-galacturonic acid, D-fructose, L-rhamnose and N-acetyl-D-galactosamine. The optimal pH and thermal stability of the purified lectin were pH 4.0-7.0 and 20-5$0^{\circ}C$, respectively.

• Journal of the Korean Society of Food Science and Nutrition
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• v.25 no.2
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• pp.240-248
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• 1996

To clarify the potentiality of sea cucumbers as dietary food, the effects of those glycoprotein on dietary proteins and physicochemical properties of those proteins were studied. Crude glycoprotein was efficiently extracted using 20mM sodium phosphate beffer(pH 7.0) and by salting out with 80% ammoniym sulfate saturation. The fractions obtained through the DEAE-cellulose ion exchange chromatography was identified as glycoprotein by Schiff's reagent and SDS polyacrylanide gel electro-phoresis. The yields of each glycoprotein from the three kinds of sea cucumbers were 0.814(red), 0.184(blue) and 0.232(black) and the molecular weights of the glycoproteins subunits were ranged from 20,000 dalton(blue and black) to 29,000 dalton(red), respectively. The electrophoretic patterns of the glycoprotein isolates were similar to each other and any significant difference in amino acid pattern was observed. Predominant arnino scids were Asx(aspartic acid and asparagine) and Glx(glutamic acid and glutamine) ; in contrast, histidine and methionine were below 2% as compared to total amino acids. water holding capacities of the glycoprotein isolates from red, blue and black cucumbers were equally 100% and emulsion activities ranged from 53% to 64%. In addition the emulsion stabilities were 7.04, 1.37 and 2.44, respectively. In vitro digestibility of some proteins(casein, SPI and squid) was decreased as increasing the level of the freeze dried sea cucumber powder and glycoprotein isolates. But squid protein was not affected.