Lectins from Marine Shells: (VIII) Characterization of the NIC Lectin Isolated and Purified from Neptunea intersculpta

서영아,전경희,정시련,Suh, Young-Ah,JeuneChung, Kyung-Hee,Chung, See-Ryun 생화학분자생물학회 1988 한국생화학회지 Vol.21 No.1

새로운 조각매물고둥렉틴인 NIC는 황화암모늄침전, 이온교환크로마토그래피 및 친화성크로마토그래피의 방법으로 분리, 정제되었다. 이는 전기영동상에서 한개의 주된 띠와 세개의 희미한 띠를 나타냈으며, specific activity는 82.16에서 85333.33 units/mg으로 증가되었다. Sephadex G-200 크로마토그래피와 SDS 전기영동결과, NIC는 분자량이 약 112,000으로 나타냈으며, 이는 분자량 약 14,000인 subunit의 octamer로 추정되었다. NIC는 2.5%의 당을 함유하는 당단백질이었으며, lactose, galactose 및 N-acetyl-D-galactosamine과 특이하게 결합하였다. 이의 아미노산조성은 산성아미노산의 함량이 많았으며, sulfur를 함유한 아마노산의 함량은 적었다. NIC, another Neptunea intersculpta lectin, was purified by salt fractionation, DEAE-cellulose 52 ion exchange chromatography and lactosyl-Sepharose 4B affinity chromatography. Through these purification procedures, specific activity of NIC was increased from 82.16 to 85333.33 units/mg. And on polyacrylamide gel electrophoresis, NIC exhibited one major and three minor bands. A molecular weight of NIC was calculated as 112,000 by Sephadex G-200 gel filtration and the subunit was determined as Mr 14,000 by sodium dodesyl sulfate polyacrylamide disc gel electrophoresis. Consequently, NIC is supposed to be an octamer composed of identical subunits of approximately Mr 14,000. NIC was a glycoprotein containing 2.5% of neutral sugar and had specificity for lactose, galactose and N-acetyl-D-galactosamine. NIC contained relatively high amounts of acidic amino acids, but the contents of sulfur containing amoino acids was very low.

해양패류로부터 렉틴성분 개발연구 : ( Ⅷ ) 조각매물고둥으로부터 분리정제된 NIC 렉틴의 특성

서영아,전경희,정시련 ( Young Ah Suh - Chae,Kyung Hee Jeune - Chung,See Ryun Chung ) 생화학분자생물학회 1988 BMB Reports Vol.21 No.1

NIC, another Neptunea intersculpta lectin, was purified by salt fractionation, DEAE-cellulose 52 ion exchange chromatography and lactosyl-Sepharose 4B affinity chromatography. Through these purification procedures, specific activity of NIC was increased from 82.16 to 85333.33 units/㎎. And on polyacrylamide gel electrophoresis, NIC exhibited one major and three minor bands. A molecular weight of NIC was calculated as 112,000 by Sephadex G-200 gel filtration and the subunit was determined as Mr 14,000 by sodium dodesyl sulfate polyacrylamide disc gel electrophoresis. Consequently, NIC is supposed to be an octamer composed of identical subunits of approximately Mr 14,000. NIC was a glycoprotein containing 2.5% of neutral sugar and had specificity for lactose, galactose and N-acetyl-D-galactosamine. NIC contained relatively high amounts of acidic amino acids, but the contents of sulfur containing amoino acids was very low.

해양패류로부터 랙틴성분 개발연구 Ⅵ 조각매물고둥으로부터 새로운 렉틴의 분리 정제 및 특성

서영아,정시련,전경희 ( Young Ah Suh - Chae,See Ryun Chung,Kyung Hee Jeune - Chung ) 생화학분자생물학회 1987 BMB Reports Vol.20 No.1

A new lectin(NIB) was obtained from the Neptunea intersculpta through the following procedure; 0.15 M NaCl extraction, (NH₄)₂SO salt fractionation, DEAE-cellulose 52, hydroxyapatite and chromatofocusing column chromatography. The hemagglutinating activity of the lectin was stable by 45℃ and was inhibited by lactose, N-acetyl-D-galactosamine and D-galactose. It`s isoelectric point was pH 3.8 and it was composed of three kinds of subunits; 68;000, 66,000 and 61,000 daltons. The lectin contained relatively high contents of acidic amino acids, but the amount of sulfur cotaining amino acids was not estimated.

Expression of c-fos, c-Ha-ras and Laminin in the Human Stomach and Colorectal Carcinoma Cell Lines

최규실,서영아,김규원,Choi, Kyu-Sil,Suh, Young-Ah,Kim, Kyo-Won 생화학분자생물학회 1991 한국생화학회지 Vol.24 No.6

한국인 위장관 종양에서 확립된 3종류의 위암 세포주(SNU-1, 5, 16)와 7종류의 대장암 세포주 (SNU-61, 67, 70, C1, C2A, C4, C5)를 대상으로 하여 암유전자 및 라미닌 유전자의 발현양상과 종양전이 능력과의 상관관계를 조사하였다. c-fos 암유전자의 경우 일차종양에서 확립된 SNU-C4와 C5에서 발현이 높게 나타나고, c-Haras 암유전자인 경우는 전이 부위에서 확립된 SNU-61 세포주에서 발현이 높으므로 전이성이 낮은 세포주에서 c-fos의 발현이 높고, 전이성이 높은 세포주에서는 c-Ha-ras의 발현이 높다는 종래의 실험결과와 어느 정도 일치함을 나타내었다. 그러나 라미니 유전자인 경우는 SNU-5, C2A, C5 등의 세포주에서 비교적 높게 발현이 되었으나 다른 세포주에서도 고르게 발현이 되고 있으므로 라미닌 유전자의 발현과 종양전이 능력과의 상관성을 SNU 세포주에서는 얻기가 어려운 것으로 추정이 된다. We have investigated a correlation between oncogene and laminin expression and metastatic aggressiveness with three stomach (SNU-1, 5, and 16) and seven colon (SNU-61, 67, 70, C1, C2A, C4, and C5) carcinoma cell lines which were estabilished from primary and metastatic tumor sites. According to Northern blot analysis and immunofluorescence staining, the expression of c fos was elevated in SNU-C4 (primary, colon) and SNU-C5 (primary, colon). The expression of c-Ha-ras was increased in SNU-61 (metastatic, colon). In malignant progression, ras oncogene expression was believed to be responsible for tumor metastasis, whereas the mRNA of c-fos was expressed mainly in the low metastatic clones. In this regard, our results were somewhat in accordance with previous reports. On the other hand, laminin mRNA was detected in all cell lines tested, and among them higher level of laminin mRNA expression was observed in SNU-5 (metastatic, stomach), SNU-C2A (primary, colon) and SNU-C5 (primary, colon). This result suggests that the tumor metastatic behavior can not be simplified to laminin gene expression and the correlation between tumor metastatic aggressiveness and laminin gene expression is more complex.

인간 위장관 종양 세포주에서 c - fos , c - Ha - ras , 그리고 라미닌의 유전자의 발현

최규실,서영아,김규원 ( Kyu Sil Choi,Young Ah Suh,Kyu Won Kim ) 생화학분자생물학회 1991 BMB Reports Vol.24 No.6

We have investigated a correlation between oncogene and laminin expression and metastatic aggressiveness with three stomach (SNU-1, 5, and 16) and seven colon (SNU-61, 67, 70, C1, C2A, C4, and C5) carcinoma cell lines which were estabilished from primary and metastatic tumor sites. According to Northern blot analysis and immunofluorescence staining, the expression of c fos was elevated in SNU-C4 (primary, colon) and SNU-C5 (primary, colon). The expression of c-Ha-ras was increased in SNU-61 (metastatic, colon). In malignant progression, ras oncogene expression was believed to be responsible for tumor metastasis, whereas the mRNA of c-fos was expressed mainly in the low metastatic clones. In this regard, our results were somewhat in accordance with previous reports. On the other hand, laminin mRNA was detected in all cell lines tested, and among them higher level of laminin mRNA expression was observed in SNU-5 (metastatic, stomach), SNU-C2A (primary, colon) and SNU-C5 (primary, colon). This result suggests that the tumor metastatic behavior can not be simplified to laminin gene expression and the correlation between tumor metastatic aggressiveness and laminin gene expression is more complex.

해양동물 렉틴 ( 제 11 보 ) : 조각매물고둥 렉틴의 림프구 분열효과 및 면역화학적 특성

전경희,서영아,소명숙,정시련 ( Kyung Hee Jeune,Young Ah Suh,Myung Suk So,See Ryun Chung ) 생화학분자생물학회 1991 BMB Reports Vol.24 No.3

Developing new substances for immunosuppressor or immunomodulator from natural products is extremely important in the present biomedicine. In this paper, we focused our efforts on the mitogenicity and immunochemical properties of the NI lectins (NIA, NIB, NIC), obtained from Neptunea intersculpta. Immunochemical techniques were employed to elucidate the structural and/or functional similarities between the NI lectins. According to these results, NIB and NIC were almost identical and the NIC proved to be homogeneous on immunoelectrophoresis as well as crossed immunoelectrophoresis. But these lectins are turned out as different molecules through the study of carbohydrate specificity. Only NIC lectin was a potent mitogenic toward murine splenic lymphocytes at a concentration ca. 6.4 ㎍/㎖.

콩과 식물에서 분리정제한 렉틴의 생물물리 화학적 연구 ( E - PHA 렉틴 )

전경희,채영흠,서영아,정시련 ( Kyung Hee Jeune - Chung,Young Heum Chae,Young Ah Suh,See Ryun Chung ) 생화학분자생물학회 1983 BMB Reports Vol.16 No.1

Salt fractionated extracts of Phaseolus vulgaris was purified successively through ion exchange chromatography on DEAE Sephadex A-50, hydroxyapatite column and EL-PHA was obtained. EL-PHA was treated with 4M urea just prior to the application into CM-Sephadex C-50 column. This column was eluted with 10 mM (pH 5.7), 20 mM (pH 6.6) and 30mM (pH 8.0) phosphate buffer, each containing 4M urea. In order to identify isolation of E-and L-PHA, erythroagglutination test with human blood A group and lymphoagglutinating test with mouse spleen lymphocytes were measured on each fraction and found three active fractions, L-PHA, having lymphoagglutinating activity only, EI-PHA, having high erythroagglutinating activity only and EII-PHA, having erythroagglutinating activity along with weak lymphoagglutinating activity. In order to characterize biophysicochemical properties of the EI-and EII-PHA, polyacrylamide disc gel electrophoresis, SDS gel electrophoresis, sugar inhibition test, carbohydrate test were performed. In polyacrylamide disc gel electrophoresis, EL-PHA was observed as three bands, EI-and EII-PHA were observed as clear single band with a very faint minor band. Molecular weight of EI-and EII-PHA were estimated at about 130,000 daltons and 125, 000 daltons, respectively by PAGE. The results of SDS-PAGE shown that EI-PHA consists of four homogenous subunits having molecular weight of 34, 000 per unit and EII-PHA consists of two different subunits with 41,000 arid 30,000 daltons. Erythroagglutination by EI-and EII-PHA was inhibited with 0.3% D-mannose and N-acetyl-D-galactosamine, but not with N-galactose, L-fucose, N-acetylneuraminic acid and N-acetyl-D-glucosamine. Sugar components of EI-and EII-PHA were recognized by anthrone test, but no spot was observed in paper chromatography. For immunochemical studies, antisera to EI-PHA were prepared by immunizing rabbit and EI-PHA formed precipitin bands against various antisera in Ouchterlony double immunodiffusion.

콩과 식물에서 분리정제한 렉틴의 생물물리 화학적 연구(E-PHA 렉틴)

전경희,채영흠,서영아,정시련,JeuneChung, Kyung-Hee,Chae, Young-Heum,Suh, Young-Ah,Chung, See-Ryun 생화학분자생물학회 1983 한국생화학회지 Vol.16 No.1

흰강남콩 (Phaseolus vulgaris)의 추출물을 DEAE Sephadex A-50과 hydroxyapatite column을 통과시켜 적혈구 및 립프구 응집효과를 갖는 EL-PHA를 얻었다. EL-PHA로부터 적혈구 응집효과만 가진 E-PHA을 분리하기 위해 EL-PHA를 4M urea로 처리한 다음 CM-Sephadex column을 통과시켜 림프구 응집력만 가진 L-PHA, 적혈구 응집력만 가진 EI-PHA, 적혈구 응집력과 약간의 림프구 응집력을 가진 ElI-PHA를 얻었다. EI-, EII-PHA의 생물물리화학적 성상을 구명코저 activity test, 전기영동, SDS-전기영동, 당류시험, 면역화학적 실험 등을 수행하였다. 전기영동에서 EL-PHA는 3개의 band를 나타내었고, EI-, EII-PHA는 1개의 뚜렷한 band를 나타내었으며, 분자량은 각각 130,000과 125,000 daltons으로 추정되었다. SDS-PAGE 결과 EI-PHA는 분자량이 약 34,000인 subunits 1종으로 나타났고, EII-PHA는 41,000과 30,000정도의 2종의 서로 다른 subunits로 나타났다. 당류에 의한 activity의 저해 효과는 0.3% D-mannose와 N-acetyl-D-galactosamine의 경우에 현저 하였고, D-galactose, L-fucose, N-acetylneuraminic acid, N-acetyl-D-glucosamine의 경우는 저해 효과가 없었다. 한편, EE-, EII-PHA 당류를 미량 함유한 당단백질임을 알 수 있었다. 면역 화학적인 연구의 하나로 immunodiffusion을 한 결과 EI-PHA는 antigen-antibody 반응을 잘 나타내 주었다. Salt fractionated extracts of Phaseolus vulgaris was purified successively through ion exchange chromatography on DEAE Sephadex A-50, hydroxyapatite column and EL-PHA was obtained. EL-PHA was treated with 4M urea just prior to the application into CM-Sephadex C-50 column. This column was eluted with 10 mM (pH 5.7), 20 mM (pH 6.6) and 30mM (pH 8.0) phosphate buffer, each containing 4M urea. In order to identify isolation of E-and L-PHA, erythroagglutination test with human blood A group and lymphoagglutinating test with mouse spleen lymphocytes were measured on each fraction and found three active fractions, L-PHA, having lymphoagglutinating activity only, EI-PHA, having high erythroagglutinating activity only and EII-PHA, having erythroagglutinating activity along with weak lymphoagglutinating activity. In order to characterize biophysicochemical properties of the EI-and EII-PHA, polyacrylamide disc gel electrophoresis, SDS gel electrophoresis, sugar inhibition test, carbohydrate test were performed. In polyacrylamide disc gel electrophoresis, EL-PHA was observed as three bands, EI-and EII-PHA were observed as clear single band with a very faint minor band. Molecular weight of EI-and EII-PHA were estimated at about 130,000 daltons and 125,000 daltons, respectively by PAGE. The results of SDS-PAGE shown that EI-PHA consists of four homogenous subunits having molecular weight of 34,000 per unit and EII-PHA consists of two different subunits with 41,000 arid 30,000 daltons. Erythroagglutination by EI-and EII-PHA was inhibited with 0.3% D-mannose and N-acetyl-D-galactosamine, but not with N-galactose, L-fucose, N-acetylneuraminic acid and N-acetyl-D-glucosamine. Sugar components of EI-and EII-PHA were recognized by anthrone test, but no spot was observed in paper chromatography. For immunochemical studies, antisera to EI-PHA were prepared by immunizing rabbit and EI-PHA formed precipitin bands against various antisera in Ouchterlony double immunodiffusion.

해양동물 눈알고둥으로부터 새로운 렉틴 성분의 분리 및 정제

소명숙(Myung Suk So),서영아(Young Ah Suh),전경희(Kyung Hee Jeune),정시련(See Ryun Chung) 대한약학회 1992 약학회지 Vol.36 No.3

The whole body extract of Lunella coronata coreensis agglutinated nonspecifically human and other animal erythrocytes. A new lectin was purified by the following procedures: 0.15 M NaCl extraction, salt fractionation, gel filtration, anionic and cationic ion exchange column chromatographies. Through these purification procedures, specific activity of LCC-I was increased from 276 to 9714.3units/mg, And on polyacrylamide gel electrophoresis, LCC-I exhibited one major band. A molecular weight of LCC-I was assumed to be 20,000 by sodium dodesyl sulfate polyacrylamide gel electrophoresis. The purified lectin was relatively stable at various pH and heat. Among the tested sugars, lactose and lactulose inhibited lectin activity at a concentration of 6.25mM, respectively.

위장관 ( 胃腸管 ) : 원발성 위선암 조직에서의 c - myc , c - H - ras , c - fos 암 유전자 , β2 - Microglobulin 및 Laminin receptor의 발현양상과 위암의 심달도 및 전이와의 관련성에 대한 연구

박병채,서진혜,강구일,허만하,서재관,이복률,서영아,김규원 ( Byung Chae Park,Jin Hye Seo,Koo Ll Kang,Man Ha Huh,Jae Kwan Suh,Bok Luel Lee,Young Ah Suh,Kyu Won Kim ) 대한소화기학회 1991 대한소화기학회지 Vol.23 No.2