• Journal of Applied Biological Chemistry
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• v.54 no.2
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• pp.84-88
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• 2011

Edestin, a major hempseed storage protein from Cheungsam, was isolated to apparent homogeneity by acid precipitation and gel filtration chromatography. The native molecular weight of purified edestin was approximately 300 kDa by Sephacryl S-300 gel filtration. Upon adding 2-mercaptoethanol, the isolated edestin of 56.7 kDa on the non-reduced sodium dodecyl sulfate polyacrylamide gel was converted into subunits, suggesting that the protein might be composed of subunits linked by disulfide bond. Cheungsam edestin was rich in essential amino acids and it has 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity. The results suggest that Cheungsam edestin might be utilized as a superior antioxidative nutrient.

• Microbiology and Biotechnology Letters
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• v.17 no.6
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• pp.587-592
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• 1989

A strain of alkalophilic Bacillus sp. YS-309 capable of producing large amount of $\beta$-galactosidase has been isolated from soil sample. Intracellular $\beta$-galactosidase was purified 6.9 folds by procedures including ammonium sulfate precipitation, DEAE-cellulose chromatography, gel-filtration, DEAE-Sephadex A-50 chromatography with over-all yield of 17.8%. The molecular weight of native enzyme was 205, 000 by HPLC, and SDS-polyacrylamide gel electrophoresis showed that the enzyme consisted of 4 identical subunits with a molecular weight of 56, 000.

• Microbiology and Biotechnology Letters
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• v.22 no.6
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• pp.627-635
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• 1994

The $\beta$-xylosidase from Penicillium sp. FX-102 was purified by 40~80% ammonium sulfate saturation, CM-Cellulose column chromatography, Sephadex G-200 gel filtration, and isoelec- tric focusing. The optimum pH and temperature for the activity of the $\beta$-xylosidase was pH 4.5 and 50$\circ$C, respectively. The enzyme was stable at the pH range of 4.5~5.5, and at 55$\circ$C for 10 min. The molecular weight of the enzyme was estimated to be about 300,000 daltons by Sephadex G-200 gel filtration and 310,000 daltons of monomer by SDS polyacrylamide gel electrophoresis. Isoelectric point of the enzyme was determined to be pH 4.4. The enzyme activity was strongly inhibited by Hg$^{2+}$, Ag$^{2+}$, n-bromosuccinimide and p-chloromercuribenzoate. Xylobiose (10 mM) was completely decomposed to xylose after 8 hrs enzyme reaction with 2 units of the $\beta$-xylosidase.

• Journal of Microbiology
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• v.34 no.1
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• pp.82-89
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• 1996

Purine nucleoside phosphorylase (PNP) was purified in Micrococcus luteus (M. luteus) using streptomycin sulfate and amomonium sulfate fractionation, three times by a Sephadex G-100 gel filtration and a DEAE-Sephadex A-50 ion exchange chromatography. The enzyme was purified 72 folds with a 11% recovery and showed a single band in a nondenaturing gel electrophoresis. The M. W. of PNP turned out to be 1.35 * 10$^{5}$ delton in G-150 gel filtration chromatography. The stability of the enzyme was increased by treatment with both substrates, MgCI$_{2}$ or CaCI$_{2}$, but not significantly kcal/mol. M. luteus PNP catalyzed the phosphorolysis of inosine, deoxyinosine, guanosine and deoxyguanosine with the Km value of 1.5 * 10$^{-3}$ M, 3.0 * 10$^{-3}$ M, 5.0 * 10$^{-4}$ M, respectively. The enzyme was reacted with adenosine, 1-methylnosine and 1-methylguanosine as substrates, which were shown to be poor substrates for mammalian enzyme.

• Microbiology and Biotechnology Letters
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• v.21 no.2
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• pp.132-138
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• 1993

The s-type pyocin was purified from the lysate of the mitomycin C-induced Pseudomonas aeruginosa 90-2-2205 cells by the other of ammonium sulfate precipitation, DEAE-Sephadex A-50 chromatography and Sephadex G-200 gel filtration. The purity was confirmed by the polyacry-lamide gel electrophoresis. The molecular weight of the purified pyocin was estimated 180, 000 by gel filtration. The pyocin was analyzed to be a complex of some polypeptides by the SDS-PAGE. The pyocin was stable by heat treatment and at pH 6-7.5 by adding 10-3% gelatin and 0.2M NaCl to the 10mM Tris-HCl buffer (pH7.5). Its killing action against the sensitive cells was assumably a single hit process.

• The Korean Journal of Zoology
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• v.37 no.1
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• pp.66-75
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• 1994

Juvenile hormone binding protein was identified in the hemolymph of fifth instar larvae and purified using column chromatography. Hemolymph was mixed with [3H] JH-III and electrophoresed on 691 NON-SDS gel, indicating that radioactivity peak appears at Rf value of 0.55. Gel filtration showed two radioactivity peaks equivalent to bound and free [3H]JH-III, respectively. JHBP was purified from hemolymph through gel filtration (Sephadex G-100), anion exchange chromatosraphv (DEAE Sepharose CL-6B), chromatofocusing chromatographv (PBE 94) and preparative electrophoresis.

• Korean Journal of Food Science and Technology
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• v.32 no.5
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• pp.1158-1167
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• 2000

To produce ${\beta}-cyclodextrin({\beta}-CD)$, a cyclodextrin glucanotransferase(CGTase) producing Aspergillus sp. CC-2-1 was isolated from soil. The enzyme was purified and its enzymological characteristics were investigated. It was found that production of CGTase reached to the maximum when the wheat bran medium containing 0.1% albumin, 2% $(NH_4)_2S_2O_8$, 2% soluble starch and 0.2% $KH_2PO_4$ was cultured for 5 days at $37^{\circ}C$. The purity of CGTase was increased by 13.14 folds after DEAE-cellulose ion exchange chromatography and Sephadex G-100, G-150 gel filtration and the specific activity was 172.14 unit/mg. Purified enzyme was confirmed as a single band by the polyacrylamide gel electrophoresis. The molecular weight of CGTase was estimated to be 27,800 by Sephadex G-100 gel filtration and SDS-polyacrylamide gel electrophoresis. The optimum pH and temperature for the CGTase activity were 9.0 and $80^{\circ}C$, respectively. The enzyme was stable in pH $8.0{\sim}11.0$ at $60{\sim}80^{\circ}C$. The activity of purified enzyme was activated by $K^+,\;Cu^{2+}$ and $Zn^{2+}$. The activity of the CGTase was inhibited by the treatment with 2,4-dinitrophenol and iodine. The result suggests that the purified enzyme has phenolic hydroxyl group of tyrosine, histidine imidazole group and terminal amino group at active site. The reaction of this enzyme followed typical Michaelis-Menten kinetics with the $K_m$ value of 18.182 g/L with the $V_{max}$ of 188.68 ${\mu}mole/min$. The activation energy for the CGTase was calculated by Arrhenius equation was 1.548 kcal/mol.

• Applied Biological Chemistry
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• v.11
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• pp.105-108
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• 1969

본(本) 실험(實驗)에 사용(使用)된 Gel로는 Sephadex G-200, G-100과 G-75로써 fractionation은 대체(大體)로 호조건에서 이루어 졌다. G-200과 G-100의 경우에 있어서는 갈색색소의 분자크기가 서로 다른 2개(個)의 Peak를 얻을 수 있었고 Colume에 투입된 시료(試料)는 완전(完全)히 회수(回收)되었다. G-75의 경우에는 갈색색소와 분자 크기가 작은 다른 물질(物質)과를 분리(分離)할 수 있었으나 약간의 시료(試料)는 Gel에 흡착되어 완전(完全)히 회수(回收)할 수 없었다. 발효실험의 결과를 종합해 보면 우선 갈색색소를 함유하늘 모든 fraction은 발효초기에 활성(活性)을 보이며 분자크기가 작은 물질의 구획(區劃)에서는 강하고도 점증되는 활성(活性)을 관찰할 수 있었다. Sephadex Gel에 의(依)한 fractionation은 첫째로 작업(作業)도중에 시료(試料)의 변질현상(變質現象)이 없다는 것과 둘째로 갈색색소를 그들의 분자크기에 따라 분획(分劃)할 수 있다는 이점(利點)들이 있다.

• 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.

• Korean Journal of Food Science and Technology
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• v.17 no.1
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• pp.1-4
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• 1985

For the systematic investigation of biochemical characteristics of Korean ginseng protein by years, protein fractions were analyzed by the techniques of polyacrylamide gel electrophoresis, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration, while the amino acid composition was studied by amino acid autoanalyzer. Results of polyacrylamide gel electrophoresis and SDS-PAGE showed a few difference in pattern and number of bands depending on the age of the root. However, the number of bands obtained from polyacrylamide gel electrophoresis and SDS-PAGE was 8 and 7 to 11, respectively. When water extracted proteins were fractionated by Sephadex G-200, the main peak among 2 peaks was collected and lyophillized. Its mol. wt. was extimated to be 43,000 by the SDS-PAGE method. In amino acid composition of water extracted protein and main fraction of gel filtration, arginine content was the highest, 47.17% in water extracted protein and 57.36% in main fraction followed by glutamic acid and asparatic acid. On the contrary, cystine and methionine contents were very low in both cases.