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Purification and Characterization of Chloroplast Fructose-1,6-bisphosphatase from Pea Leaves
부성희,한태룡,Bhoo, Seong-Hee,Hahn, Tae-Ryong 생화학분자생물학회 1989 한국생화학회지 Vol.22 No.4
엽록체 fructose-1,6-bisphosphatase를 완두 잎으로부터 1,300배 정제하였다. 정제된 효소는 거의 순수하였고 SDS polyacrylamide 전기영동하여 얻은 monomer의 분자량은 약 40,000이었다. 효소는 pH 7.5에서 활성이 없었으나 dithiothreitol 혹은 알카리 pH에서 활성화가 되었다. 이 결과는 분리된 fructose-1,6-bisphosphatase가 엽록체로부터 얻어진 것임을 나타낸다. Fructose-1,6-phosphate와 $Mg^{++}$에 대한 효소 포회곡선은 Hill 상수 2.6 및 3.1을 가진 sigmoidal한 모양을 보여주는데 이는 효소가 multimer로 구성된 allosteric 한 것임을 알 수 있다. 최고 활성의 반에 요구되는 기질농도는 $40{\mu}M$로서 이는 시금치 엽록체 효소에 대한 $80{\mu}M$값 (Zimmermann et al., 1976)보다 작았다. Chloroplast fructose-1,6-bisphosphatase was purified 1,300 fold from pea leaves. The purified enzyme appeared homogeneous and the approximate molecular weight of the monomer was 40,000, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme was inactive at pH 7.5, while it was activated by dithiothreitol or alkaline pH, indicating that the purified fructose-1,6-bisphosphatase was originated from chloroplast. The enzyme saturation curves with fructose-1,6-bisphosphate and $Mg^{++}$ show sigmoidal shapes with almost same Hill coefficients (2.6 and 3.1, respectively), suggesting that the enzyme is composed of multimer. The substrate concentration required for half-maximal activity was $40{\mu}M$ which is a comparable value $(80{\mu}M)$ for spinach chloroplast fructose bisphosphatase (Zimmermann et al., 1976).
Roles of Ki67 in Breast Cancer - Important for Management?
Yip, CH,Bhoo-Pathy, N,Daniel, JM,Foo, YC,Mohamed, AK,Abdullah, MM,Ng, YS,Yap, BK,Pathmanathan, R Asian Pacific Journal of Cancer Prevention 2016 Asian Pacific journal of cancer prevention Vol.17 No.3
Background: The three standard biomarkers used in breast cancer are the estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2). The Ki-67 index, a proliferative marker, has been shown to be associated with a poorer outcome, and despite absence of standardization of pathological assessment, is widely used for therapy decision making. We aim to study the role of the Ki-67 index in a group of Asian women with breast cancer. Materials and Methods: A total of 450 women newly diagnosed with Stage 1 to 3 invasive breast cancer in a single centre from July 2013 to Dec 2014 were included in this study. Univariable and multivariable logistic regression was used to determine the association between Ki-67 (positive defined as 14% and above) and age, ethnicity, grade, mitotic index, ER, PR, HER2, lymph node status and size. All analyses were performed using SPSS Version 22. Results: In univariable analysis, Ki -67 index was associated with younger age, higher grade, ER and PR negativity, HER2 positivity, high mitotic index and positive lymph nodes. However on multivariable analysis only tumour size, grade, PR and HER2 remained significant. Out of 102 stage 1 patients who had ER positive/PR positive/HER2 negative tumours and non-grade 3, only 5 (4.9%) had a positive Ki-67 index and may have been offered chemotherapy. However, it is interesting to note that none of these patients received chemotherapy. Conclusions: Information on Ki67 would have potentially changed management in an insignificant proportion of patients with stage 1 breast cancer.
白南豪,朴富植 서울대학교 1972 서울대학교 論文集 Vol.22 No.-
In an acidic solution caffeine reacts with phosphomolybdenic acid to form a yellow precipitate, which can be extracted with diethyl ether and then readily reduced by 5% stannous chloride solution to form a molybden blue, showing the absorbance maximum at 623mμ. By adjusting the concentration of hydrochloric acid caffeine phosphomolybdate can be extracted and separated from the unreacted polyheteroacid, and caffeine is measured quantitatively by the spectrophotometric determination of molybden blue.
Biochemical characterization of a regiospecific flavonoid 3'-O-methyltransferase from orange
Park Hye Lin,Bhoo Seong Hee,Lee Sang-Won,Cho Man-Ho 한국응용생명화학회 2024 Applied Biological Chemistry (Appl Biol Chem) Vol.67 No.-
Citrus plants have diverse methoxyflavonoids including, chrysoeriol, isosakuranetin, and nobiletin. In plants, O-methyltransferases (OMTs) participate in the methylation of a vast array of secondary metabolites, including flavonoids, phenylpropanoids, and alkaloids. To identify functional OMTs involved in the formation of methoxyflavonoids, orange (Citrus sinensis) OMT (CsOMT) genes were retrieved from the Citrus Genome Database. The phylogenetic relationships with functional OMTs suggested that three CsOMTs, CsOMT15, CsOMT16, and CsOMT30, are possible candidates for flavonoid OMTs (FOMTs). These CsOMTs were heterologously expressed in Escherichia coli, and their OMT activity was examined with flavonoid substrates. Of the examined CsOMTs, CsOMT16 catalyzed the regiospecific 3’-O-methylation of flavonoids to the respective 3’-methoxyflavonoids. A kinetic study demonstrated that CsOMT16 accepts diverse flavonoids as a substrate with a comparable preference. The flavonoids eriodictyol, luteolin, and quercetin were efficiently converted to homoeriodictyol, chrysoeriol, and isorhamnetin by CsOMT16- transformed E. coli cells, respectively. These findings suggest that CsOMT16 contributes to the methoxyflavonoid formation in orange and is applicable to the biotechnological production of 3’-methoxyflavonoids.
완두엽록체 fructose - 1 , 6 - bisphosphatase 의 분리정제 및 특성
부성희,한태룡 ( Seong Hee Bhoo,Tae Ryong Hahn ) 생화학분자생물학회 1989 BMB Reports Vol.22 No.4
Chloroplast fructose-1,6-bisphosphatase was purified 1,300 fold from pea leaves. The purified enzyme appeared homogeneous and the approximate molecular weight of the monomer was 40,000, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme was inactive at pH 7.5, while it was activated by dithiothreitol or alkaline pH, indicating that the purified fructose-1,6-bisphosphatase was originated from chloroplast. The enzyme saturation curves with fructose-1,6-bisphosphate and Mg^(++) show sigmoidal shapes with almost same Hill coefficients (2.6 and 3.1, respectively), suggesting that the enzyme is composed of multimer. The substrate concentration required for half-maximal activity was 40 μM which is a comparable value (80μM) for spinach chloroplast fructose bisphosphatase (Zimmermann et al., 1976).
Kyungha Lee,Seong Hee Bhoo,Sang‑Won Lee,Man‑Ho Cho 한국응용생명화학회 2024 Applied Biological Chemistry (Appl Biol Chem) Vol.67 No.-
Rhododendrons produce a variety of methoxyflavonoids, including rarely found 3-methoxyflavonoids and 5-methoxyflavonoids. It was thus suggested that they have a series of regiospecific flavonoid O-methyltransferases (FOMTs). The 18 Class II O-methyltransferase (OMT) genes were retrieved from the Rhododendron delavayi genome, designating them as RdOMTs. A comprehensive biochemical characterization of RdOMTs was performed to identify functional FOMTs. The FOMT activity of recombinant RdOMTs was assayed with flavonoid substrates of different subclasses. Among the examined RdOMTs, RdOMT3, RdOMT10, and RdOMT12 showed FOMT activity for diverse flavonoids. In particular, RdOMT3 consumed only flavonols as a substrate. Structural analyses of the methylated products demonstrated that RdOMT3, RdOMT10, and RdOMT12 catalyze regiospecific methylation of flavonoids at the 3’/5’-, 3-, and 4’-hydroxyl groups, respectively. Their broad substrate spectrum and different regiospecificity suggest that these RdOMTs contribute to the formation of complex methoxyflavonoids in R. delavayi. Bioconversion of flavonoids using E. coli harboring each RdOMT demonstrated that RdOMT3, RdOMT10, and RdOMT12 are useful tools for the biotechnological production of valuable methoxyflavonoids, including the rarely found 3-methoxyflavonoids. Rhododendrons produce a variety of methoxyflavonoids, including rarely found 3-methoxyflavonoids and 5-methoxyflavonoids. It was thus suggested that they have a series of regiospecific flavonoid O -methyltransferases (FOMTs). The 18 Class II O -methyltransferase (OMT) genes were retrieved from the Rhododendron delavayi genome, designating them as RdOMTs. A comprehensive biochemical characterization of RdOMTs was performed to identify functional FOMTs. The FOMT activity of recombinant RdOMTs was assayed with flavonoid substrates of different subclasses. Among the examined RdOMTs, RdOMT3, RdOMT10, and RdOMT12 showed FOMT activity for diverse flavonoids. In particular, RdOMT3 consumed only flavonols as a substrate. Structural analyses of the methylated products demonstrated that RdOMT3, RdOMT10, and RdOMT12 catalyze regiospecific methylation of flavonoids at the 3'/5'-, 3-, and 4'-hydroxyl groups, respectively. Their broad substrate spectrum and different regiospecificity suggest that these RdOMTs contribute to the formation of complex methoxyflavonoids in R. delavayi . Bioconversion of flavonoids using E. coli harboring each RdOMT demonstrated that RdOMT3, RdOMT10, and RdOMT12 are useful tools for the biotechnological production of valuable methoxyflavonoids, including the rarely found 3-methoxyflavonoids.