• Journal of Embryo Transfer
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• v.13 no.3
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• pp.301-312
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• 1998

To efficiently separate a protein stimulating sperm swim-up migration and movement from follicular proteins, the effect of paper chromatography and liquid chromatography with reverse phase column and superose column on protein separation was examined. And the results obtained were as follows; 1. The band component that was separated with paper chromatography stimulated sperm migration and movement depending on its additional levels. Especially, band I component significantly increased sperm migration. But, all components of bands 1, 2 and 3 showed lower sperm migration and movement, compared to follicular fluid at the same additional level. 2. Among the components separated from follicular protein of 2~5mm follicles with reverse phase column ($\mu$RPC), components at retention time (RT) of 3.33, 7.00, 13.87, and 16.6A minutes stimulated sperm migration within a limited range. 3. All components separated from follicular protein of 10mm follicles with $\mu$RPC column didn't stimulate sperm migration and movement. 4. Among the components separated from follicular protein of 2~5m follicles with superose column, components at retention volume (RV) of 1.35 and 0.82 ml significantly stimulated sperm migration and movement. In conclusion, protein components stimulating sperm migration and movement were efficiently separated with superose column in Smart system. Especially, components of RV 1.35 and RV0.82 stimulated sperm swim-up separation.

• Journal of Embryo Transfer
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• v.14 no.1
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• pp.47-57
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• 1999

Among proteins separated from methanol extract of follicular fluid with superose column, the components inducing sperm swim-up separation through sucrose layer were analysed with superose column in Smart system and SDS-PAGE. And the results obtained were as follows; The fractions of retention volume (RV) 0.83ml and RV 1.36ml separated with superose column should stimulate sperm migration and movement. However, RV 0.83 fraction was consisted of complex materials containing RV 1.36 component. RV 1.36 fraction contained a BSA analogue of 67 kilodaltons (Kd) and showed identical peak pattern with BSA fraction V. In conclusion, the protein of 67 Kd in follicular fluid should stimulate sperm migration and movement.

• YAKHAK HOEJI
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• v.41 no.5
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• pp.638-646
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• 1997

The steroid 9${alpha}$-hydroxylase activity has been detected in cytosol fraction, $100,00{\times}g$ supernatant of cell free extract of Mycobacterium fortuitum. The activity was not linear with protein concentration in the assay suggesting 9${alpha}$-hydroxylase is a multicomponent enzyme. The 9${alpha}$-hydroxylase system was partially purified through fractional saturation of ammonium sulfate, strong anion exchange (Mono Q) column chromatography, gel filtration (Superose 12) column chromatography, and testosterone affinity gel chromatography. Ammonium sulfate 50~60% saturated fraction of the cytosol gave 9${alpha}$-hydroxylase activity. For further purification, the half-saturated ammonium sulfate fraction was applied to Mono Q, Superose 12, or affinity gel column. The purification factors of 9${alpha}$-hydroxylase containing fraction after Mono Q, Superose 12, and affinity gel chromatography was 13, 11, and 17 respectively.

• Proceedings of the KSAR Conference
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• 2002.06a
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• pp.12-12
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• 2002

난포액은 난자의 성숙을 억제하는 성분을 함유하고 있으나, 억제성분의 화학적 성질은 연구자에 따라 다르게 보고되고 있다. 따라서 본 연구는 난자의 성축과 관련하여 제l극체의 형성을 억제하는 단백성 성분을 화학적으로 규명하기 위하여 실시하였다. 난포액 함유 난자성숙 억제 인자를 분리하기 위하여 난포액을 methanol로 추출한 다음 Superose 12 및 Superdex column 을 이 용하여 gel filtration 을 실시하였다. 회수한 Superdex 분절을 PITC로 처리한 다음, Amino Acid Analysis column을 이용, 억제 인자를 동정하였다. (중략)

• Journal of Embryo Transfer
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• v.15 no.3
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• pp.237-245
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• 2000

This study was carried out to elucidate whether sperm contain a factor inducing second polar body extrusion and to search for an effective collection method of the sperm factor Thus, sperm extract, dialyzed sperm-extract or liquid chromatographic fractions of sperm extract was microinjected into ovulated oocytes. And the microinjected oocytes were incubated for 24 hours to investigate about the extrusion of second polar body. The results obtained were as follows; 1. Sperm extract significantly increased the second polar body extrusion. 2. Sperm extract showed five major fractions at retention volumes (RVs) 1.25, 1.37, 1.84, 2.10 and 2.67ml after separation with Superose 12 column. These sperm extract fractions did not significantly increase the second polar body extrusion. 3. Dialyzed sperm-extract significantly increased the second polar body extrusion 4. Dialyzed sperm-extract showed three maior fractions at RVs 1.88, 2.14 and 2.77ml after separation with Superose 12 column. Of these fractions, the fraction RV2.14 significantly increased the second polar body extrusion. In conclusion, sperm extract contained a factor inducing the second polar body extrusion and the factor was contained largely in fraction RV2.14 after dialysis and liquid chromatographic fractionation of sperm extract.

• Microbiology and Biotechnology Letters
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• v.20 no.4
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• pp.401-408
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• 1992

The extracellular cholesterol oxidase produced from a soil microorganism HSL613 was purified and partially characterized. Through a series of purification procedures including concentration with CH2 concentrator, DEAE-cellulose column chromatography and gel filtration on Superose12, the purified enzyme was shown to have a specific activity of 108 units/mg protein giving 30.8-fold purification and final yield of 66%. The molecular weight of the enzyme was estimated to be 59,500 daltons by SDS-PAGE. The optimum temperature and pH for this enzyme were $50^{\circ}C$ and 6.0, respectively. The activity of the purified cholesterol oxidase was inhibited by $Ag^{2+}$, $Hg^{2+}$ and SDS.

• Biomedical Science Letters
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• v.2 no.1
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• pp.21-30
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• 1996

MAP kinases are a family of serine/threonine specific protein kinases becoming activated in response to different proliferative stimuli by phosphorylation at both threonine and tyrosine residue. Present study shows that MAP kinase was purified from P388 murine leukema cells by SP sephadex C-50, phenyl superose and Mono Q column chromatography and identified with anti-ERKl antibody by western blotting. Immnublotting analysis to the crude extract of P388 cell lysate shows 44 kD and other minor bands but partial purified fraction eluted from phenyl supherose column have 44kD and 66 kD isoform. Subcloned GST-fusion protein from N-terminal of $p56^{kk}$ was tested as a substrate for MAP kinase phosphorylation. It was showed that the wild type and mutant forms(S42A) were fully phosporylated by purified MAP kinase fraction as com-pare with the other mutant form(S59A). This finding suggest that those GST-fusion proteins may be used as substrate for the in vitro test of MAP kinase.

• Korean Journal of Animal Reproduction
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• v.26 no.2
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• pp.165-171
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• 2002

Follicular fluid (FF) contains an oocyte maturation inhibitor with unknown chemical properties. This study was carried out to chemically define the factor(s) inhibiting cumulus cell denudation (CD) and first polar body formation (PBF). Porcine FF (PFF) was extracted with methanol and the extract was serially separated using gel filtration on Superose 12 and Superdex columns. A Superdex fraction was derived with PITC and analyzed with an amino acid analysis column. The results obtained are as follows; PFF had an activity inhibiting both CD and PBF of porcine primary oocytes. Superdex fractions RV2.11 prepared from PFF exhibited an activity inhibiting CD and PBF. By amino acid analysis, the fraction RV2.11 appeared to be proline having the same activity inhibiting CD and PBF. In conclusion, PFF had oocyte maturation inhibitors, of which proline should inhibit CD and PBF.

• Journal of Dairy Science and Biotechnology
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• v.15 no.1
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• pp.1-9
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• 1997

This study was conducted to efficiently separate the Ig from Korean native cattle colostrum and to utilize them as an immunogen for the production of antibodies aginst rabbit. The results obtained were as follows : 1. About 84% of Ig G could be separated from Korean native cattle colostrum by·gel filtration using Superose 12 column on HPLC. The separation profile of Korean native cattle colostral immunoglobulin was similar that of Holstein colostral Ig. 2. Separation of Korean native cattle colostral Ig by anion exchange chromatography using Mono Q column on HPLC was poor resolution chromatographic pattern. 3. Hi-Trap Protein G column showed better results than the Protein A Sepharose CL-4B column in the Ig G binding capacity from Korean native cattle colostral Ig. 4. Protein G Sepharose Fast Flow system resulted in higher Ig g binding capacity as the industrial size scale-up approach. 5. Sufficient titer reaction of antibody to Korean native cattle colostral Ig G was confirmed by ELISA.

• 한국생물공학회:학술대회논문집
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• 2000.04a
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• pp.485-488
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• 2000

Methyl mercaptan oxidase was isolated and purified from Thiobacillus thioparus TK-m for the detection of mercaptans. The procedure of purification involved DEAE-Sephacel and Superose 12 column chromatographies with recovery yields of 47.5 and 48.5 %, and specific activity of 374 and 1240.8 units/mg-protein, respectively. The molecular weight of purified methyl mercaptan oxidase was determined to be 66.1 kDa by SDS-PAGE. Optimum temperature for activity was observed at $55\;^{circ}C$. This enzyme was activated by $(NH_4)_2SO_4$ and NaCl and inhibited by $NH_4Cl$.