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Expression and pH-dependence of the Photosystem II Subunit S from Arabidopsis thaliana
정미숙,황은영,Gyoung-Ean Jin,박소영,Ismayil S. Zulfugarov,문용환,이춘환,장세복 대한화학회 2010 Bulletin of the Korean Chemical Society Vol.31 No.6
Photosynthesis uses light energy to drive the oxidation of water at an oxygen-evolving catalytic site within photosystem II (PSII). Chlorophyll binding by the photosystem II subunit S protein, PsbS, was found to be necessary for energy-dependent quenching (qE), the major energy-dependent component of non-photochemical quenching (NPQ) in Arabidopsis thaliana. It is proposed that PsbS acts as a trigger of the conformational change that leads to the establishment of nonphotochemical quenching. However, the exact structure and function of PsbS in PSII are still unknown. Here, we clone and express the recombinant PsbS gene from Arabidopsis thaliana in E. coli and purify the resulting homogeneous protein. We used various biochemical and biophysical techniques to elucidate PsbS structure and function, including circular dichroism (CD), fluorescence, and DSC. The protein shows optimal stability at 4 oC and pH 7.5. The CD spectra of PsbS show that the conformational changes of the protein were strongly dependent on pH conditions. The CD curve for PsbS at pH 10.5 curve had the deepest negative peak and the peak of PsbS at pH 4.5 was the least negative. The fluorescence emission spectrum of the purified PsbS protein was also measured, and the λmax was found to be at 328 nm. PsbS revealed some structural changes under varying temperature and oxygen gas condition.
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엄수나(Suna Um),진경언(Gyoung-ean Jin),박계원(Kye Won Park),유영복(Young-bok Yu),박기문(Ki-Moon Park) 한국식품과학회 2010 한국식품과학회지 Vol.42 No.1
일반 느타리 13품종과 색상 느타리 5품종을 사용하여 아미노산 및 polyphenol, β-glucan 함량을 분석하고, 생리활성으로 항산화 및 항암, 항고혈압, 항혈전, 항당뇨, 항염활성을 측정하였다. 느타리버섯 18종의 아미노산 분석결과 전반적으로 감칠맛을 내는 glutamic acid 함량이 비교적 많이 함유되어 있었고, 필수아미노산 성분도 고르게 분포되어 있었다. Polyphenol 함량에서는 전 품종에서 20 mg% 함량이상을 나타냈으며, 노랑느타리(R)가 39.13±0.82 mg%로 가장 높았다. β-glucan 함량은 노랑느타리(R)에서 37.67±0.22%로 가장 높았으며, 그 외에 원형1(C), 장안PK(A)에서 각각 28.75±0.61%, 27.95±0.33%의 순으로 나타났다. 전자공여능에서는 노랑느타리(R) 버섯의 DPPH IC50값이 2.93±0.44 mg/mL로 가장 낮아 항산화 활성이 가장 우수한 것으로 나타났으며, 세포독성 실험에서는 노랑느타리(R) 에탄올 추출물 1% 처리시 신장 암세포에 대해 36.90%의 세포 억제율을 보였다. ACE 저해활성의 경우 노랑느타리(R) 에탄올 추출물 1%농도에서 60.5±0.2%의 저해율이 측정되었고, 흑평(B) 56.7±1.1%, 여름(H) 52.4±1.3% 수준으로 나타났다. 항혈전 활성에서는 3%농도에서 흑평(B)과 삼복(G)을 제외한 나머지 느타리버섯 에탄올 추출물에서 50%이상의 용해 활성을 보였으며 노랑느타리(R)에서 거의 plasmin과 동등한 활성을 나타냈다. 항당뇨 활성에서는 노랑느타리(R)의 경우 50.5±0.8%의 비교적 높은 효소저해율이 측정되었고, 항염활성에서는 노랑느타리(R)에서 68.4±0.3%의 억제율이 측정되었다. 이상의 결과로 일반 느타리 13 품종과 육종 재배된 색상 느타리 5품종 중 노랑느타리(R)가 가장 우수한 생리활성을 나타내 향후 기능성 소재로의 활용가능성이 기대되었다. In this study, the anti-oxidant, anti-tumorigenic, anti-hypertensive, anti-thrombic, anti-diabetic, and antiinflammatory properties of 18 different species of genus Pleurotus were investigated. In addition, the amino acid, β-glucan, and polyphenol content were also measured. All species contained more than 20 mg% of polyphenol with the highest contents found in Pleurotus cornucopiae var. citrinopileatus (yellow pleurotus) (39.13±0.82 mg%). The β-glucan contents was also the highest in yellow Pleurotus (37.67±0.22%) followed by Won-Hyeong1 (C, 28.75±0.61%) and Jang-an PK (A, 27.95±0.33%). The yellow Pleurotus exhibited the highest antioxidant activity as assessed by the DPPH scavenging rate with an IC50 value of 2.94±0.44 mg/mL. Ethanol extracts from the yellow Pleurotus treated at 1% concentration showed cytotoxic activity up to 36.9% in the human embryonic kidney 293T cell lines. The yellow Pleurotus also showed the highest inhibitory effects on ACE activity (60.52±0.2%). Finally, the yellow Pleurotus exhibited anti-diabetic and antiinflammatory properties as shown by inhibition of α-amyloglucosidase activity (50.5±0.8%) and nitric oxide production (68.4±0.3%). Taken together, our data indicate the yellow pleurotus is a promising functional food ingredients.
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The Binding of Human CLIC1 with SEDL and Its Characterization in vitro
박정순,Kyoung Mi Lee,정미숙,Gyoung-Ean Jin,장세복 대한화학회 2007 Bulletin of the Korean Chemical Society Vol.28 No.4
Full-length chloride intracellular channel protein 1 (CLIC1) is a member of the family of proteins related to bovine chloride intracellular channel p64. Mutations in the SEDL gene cause spondyloepiphyseal dysplasia tarda (SEDT), a rare X-linked chondrodysplasia. The link between the intracellular chloride channels and SEDL is an important step toward understanding their functional interplay. In the present study, CLIC1 protein was subcloned into the pGEX-KG vector and overexpressed in XL-1 blue cells. We developed a large-scale expression system composed of glutathione S-transferase (GST) fused with a 240-amino-acid CLIC1 protein in Escherichia coli. The soluble CLIC1 protein was successfully purified to homogeneity, and its purity, identity, activity and conformation were determined using SDS-PAGE, MALDI-MS, biophotometer and circular dichroism spectroscopic studies. The binding of both CLIC1 and SEDL proteins in vitro was detected by BIAcore biosensor and fluorescence measurements.
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Expression and pH-dependence of the Photosystem II Subunit S from Arabidopsis thaliana
Jeong, Mi-Suk,Hwang, Eun-Young,Jin, Gyoung-Ean,Park, So-Young,Zulfugarov, Ismayil S.,Moon, Yong-Hwan,Lee, Choon-Hwan,Jang, Se-Bok Korean Chemical Society 2010 Bulletin of the Korean Chemical Society Vol.31 No.6
Photosynthesis uses light energy to drive the oxidation of water at an oxygen-evolving catalytic site within photosystem II (PSII). Chlorophyll binding by the photosystem II subunit S protein, PsbS, was found to be necessary for energy-dependent quenching (qE), the major energy-dependent component of non-photochemical quenching (NPQ) in Arabidopsis thaliana. It is proposed that PsbS acts as a trigger of the conformational change that leads to the establishment of nonphotochemical quenching. However, the exact structure and function of PsbS in PSII are still unknown. Here, we clone and express the recombinant PsbS gene from Arabidopsis thaliana in E. coli and purify the resulting homogeneous protein. We used various biochemical and biophysical techniques to elucidate PsbS structure and function, including circular dichroism (CD), fluorescence, and DSC. The protein shows optimal stability at $4^{\circ}C$ and pH 7.5. The CD spectra of PsbS show that the conformational changes of the protein were strongly dependent on pH conditions. The CD curve for PsbS at pH 10.5 curve had the deepest negative peak and the peak of PsbS at pH 4.5 was the least negative. The fluorescence emission spectrum of the purified PsbS protein was also measured, and the ${\lambda}_{max}$ was found to be at 328 nm. PsbS revealed some structural changes under varying temperature and oxygen gas condition.
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Physiological activity and nutritional composition of Pleurotus species
Se-hyun Jo,Suna-Um,Gyoung-ean Jin,Kye Won Park,Young-bok Yu,Ki-Moon Park 한국버섯학회 2010 한국버섯학회지 Vol.8 No.4
In this study, the anti-oxidant, anti-tumorigenic, anti-hypertensive, anti-thrombic, anti-diabetic, and anti- inflammatory properties of 18 different species of genus Pleurotus were investigated. In addition, the amino acid, β-glucan, and polyphenol contents were also measured. The β-glucan and polyphenol contents were the highest in Pleurotus cornucopiae var. citrinopileatus (yellow Pleurotus) of all species. The yellow Pleurotus also exhibited the highest physiological activity as assessed by the DPPH IC, Cytotoxic activity, ACE activity and so on. To confirm the mechanism for the physiological activity of the yellow Pleurotus, we performed further examinations within ICR mice. The yellow Pleurotus reduced glucose, triglyceride and total cholesterol levels in the ICR mice blood for 4 weeks after feeding, and also significantly lowered both GOT and GPT levels. Taken together, our data indicates the yellow pleurotus is a promising functional food ingredient.
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The Binding of Human CLIC1 with SEDL and Its Characterization in vitro
Park, Jeong-Soon,Lee, Kyoung-Mi,Jeong, Mi-Suk,Jin, Gyoung-Ean,Jang, Se-Bok Korean Chemical Society 2007 Bulletin of the Korean Chemical Society Vol.28 No.4
Full-length chloride intracellular channel protein 1 (CLIC1) is a member of the family of proteins related to bovine chloride intracellular channel p64. Mutations in the SEDL gene cause spondyloepiphyseal dysplasia tarda (SEDT), a rare X-linked chondrodysplasia. The link between the intracellular chloride channels and SEDL is an important step toward understanding their functional interplay. In the present study, CLIC1 protein was subcloned into the pGEX-KG vector and overexpressed in XL-1 blue cells. We developed a large-scale expression system composed of glutathione S-transferase (GST) fused with a 240-amino-acid CLIC1 protein in Escherichia coli. The soluble CLIC1 protein was successfully purified to homogeneity, and its purity, identity, activity and conformation were determined using SDS-PAGE, MALDI-MS, biophotometer and circular dichroism spectroscopic studies. The binding of both CLIC1 and SEDL proteins in vitro was detected by BIAcore biosensor and fluorescence measurements.
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