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Chong, Chom-Kyu;Chang, Soo-Ik;Choi, Jung-Do
- BMB Reports
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- v.30 no.4
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- pp.274-279
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- 1997
Acetolactate synthase (ALS) catalyzes the first common step in the biosynthesis of branchedchain amino acids, valine, leucine, and isoleucine. ALS is the target site for several structually diverse classes of herbicides including sulfonylureas, imidazolinones. and triazolopyrimidines. We have purified ALS from etiolated barley shoots to homogeneity. The five major purification steps are ammonium sulfate fractionation, DEAE anion exchange, hydroxylapatite, Bio-Gel A gel filtration, and low pressure Mono-Q chrornatoqraphy. Approximately 170-fold purification was achieved and the yield was 0.45% of initial activity in the crude extract. Both SDS-PAGE and Western blot analysis showed a single polypeptide of ALS with an apparent molecular mass of 64 kDa. The result of nondenaturing gel electrophoresis with activity staining indicated that the molecular mass of its native form is approximately 225 to 250 kDa. The values of $K_m$ for pyruvate. pl. and optimum pH of ALS were determined to be 2.0 mM, 5.2. and 7.0. respectively Feedback inhibition studies showed that ALS is more susceptible to leucine than valine. And $IC_{50}$ value of Cadre, a class of irnidazolinones, is about $1.5\mu{M}$ for ALS.
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Park, Yu-Geun;Song, Jooyoung;Kim, Yongae
- Journal of the Korean Magnetic Resonance Society
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- v.16 no.2
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- pp.147-161
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- 2012
Human melanocortin-4 receptor (hMC4R) among MC-Rs, expressed in the brain, is in charge of the control on energy homeostasis and food intake. The structure and function of human MC4R have been studied to understand their essential function and roles. To investigate the structure and function, it is necessary to prepare sufficient amounts of proteins. However, their expression and purification is demanding and time-consuming due to their innate insoluble and toxic properties. The heterozygous mutations of hMC4R, exchange of Asp 90 to Asn located in second transmembrane, cause severe obesity in human. To obtain purified hMC4R wt-TM2 for structural studies, it was first over-expressed and purified by fast protein liquid chromatography (FPLC) and then solution NMR studies were performed to get high-resolution spectra. In here, we established optimized purification scheme to get more purified target peptide.
https://doi.org/10.6564/JKMRS.2012.16.2.147 인용 PDF KSCI

이양봉;신의철;김병철;양지영;장영진
- Journal of the Korean Society of Food Science and Nutrition
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- v.27 no.5
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- pp.808-812
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- 1998
Melania snail(Semisulcopira bensoni) is used as ingredient in Korean traditional soup and nutritional foods. Generally, lipoxygenase in several food products may produce off-flavors during their processing and storage. Therefore, the inactivation of lipoxygenase is required to make the better extracts from Melania sanil. Also, the quality on freshness of Melania snail may be evaluated by lipoxygenase activity. The lipoxygenae activity was the highest at 40~60% saturation among several concentrations in salting-ouot saturated solution of ammonium sulfate. The partial purification of lipoxygenase was successfully obtained by Sephacryl S-200 gel chromatography. The first peak among three peaks for protein determination showed the highest activity of lipoxygenase in 13~16 fractions among 100 fractions. The highest peak of lipoxygenase activity by ion exchange chromatography was shown at 0.1M NaCl. In the purification step, the specific activity was 20.8U/mg and activity yield was 19.8%. The optimum pH and temperature were pH6.0~8.0 and 3$0^{\circ}C$, respectively. Molecular weight of the lipoxygenase was estimated about 35kDa by SDS-PAGE.
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