条斑紫菜高纯度藻红蛋白规模制备及光学性质研究

生物技术通报 ›› 2013, Vol. 0 ›› Issue (3): 192-198.

• 研究报告 • 上一篇

条斑紫菜高纯度藻红蛋白规模制备及光学性质研究

郭子叶¹,蔡春尔^1,2,李春霞¹,耿中雷¹,贾睿^1,2,何培民^1,2

1. 上海海洋大学水产与生命学院，上海 201306 ；2. 上海海洋大学海洋科学研究院，上海 201306

收稿日期:2012-09-23 修回日期:2013-03-21 出版日期:2013-03-20 发布日期:2013-03-21
作者简介:郭子叶，女，硕士研究生，研究方向：海藻生物活性物质；E-mail ：luckyguoziye@gmail.com
基金资助:
国家科技支撑计划课题（2012BAC07B03），上海市教委重点学科项目（S30701）

Pilot Preparation and Optical Properties of Highly Purified Phycoerythrin from Porphyra yezoensis

Guo Ziye¹, Cai Chuner^{¹, 2}, Li Chunxia¹, Geng Zhonglei¹, Jia Rui^{¹, 2}, He Peimin^{¹, 2}

1. College of Fisheries and Life Science,Shanghai Ocean University,Shanghai 201306 ；2. Institute of Marine Science,Shanghai Ocean University,Shanghai 201306）

Received:2012-09-23 Revised:2013-03-21 Published:2013-03-20 Online:2013-03-21

摘要/Abstract

摘要： 旨在大量制备条斑紫菜高纯度藻红蛋白并研究其光学性质。以江苏吕泗条斑紫菜为原料，采用“溶胀+ 组织捣碎” 法破碎2 kg 冻干条斑紫菜叶状体细胞，经多次硫酸铵盐析和一次羟基磷灰石层析后获得纯度＞ 3.20 的藻红蛋白的产率为0.184%，再经一次同样层析可获得纯度＞ 4.50 的藻红蛋白的产率为0.109%。蛋白纯度已通过吸收/ 荧光激发光谱，十二烷基硫酸钠- 聚丙烯酰胺凝胶电泳（SDS-PAGE）/ 非变性聚丙烯酰胺凝胶电泳（Native-PAGE）的考马斯亮蓝/ 醋酸锌双染验证。该蛋白摩尔消光系数经Folin-酚法和Bradford 法测定分别为1.79 mL/（mg?cm）和1.73 mL/（mg?cm），20℃时在505 nm 处的荧光量子产率为0.90，在日光灯、发光二极管、碘钨灯和激光4 种光源下的光漂白效果以激光最显著，240 μg/mL 藻红蛋白10 min 可接近碘钨灯辐照240 min 的光漂白率。

关键词: 条斑紫菜, 藻红蛋白, 纯化, 摩尔消光系数, 荧光量子产率, 光漂白

Abstract: Pilot preparation were tried to obtain highly purified phycoerythrin（PE）in abundance from P. yezoensis in Lvsi city, Jiangsu Province and to study its optical properties. 2 kg freeze-dried P. yezoensis thallus was broken using “swelling & smash” method. Further purification was performed by means of ammonium sulphate co-precipitation（ASCP）and hydroxyapatite（HA）column chromatogram and the purity of PE achieved 1.67（A565/A280）, corresponding to yield of 0.184%, The purity of PE could be improved beyond 4.5 with a yield of 0.109% if HAP chromatography repeated. PE was validated by absorption/fluorescence spectra and SDS-PAGE/Native-PAGE stained with coomassie brilliant blue/zinc acetate for its purity. Molar extinction coefficient（MEC）of PE were 1.79mL/（mg?cm）and 1.73 mL/（mg?cm） determined by folin-phenol method and Bradford method, respectively. Fluorescence quantum yield of PE was 0.90 at 505 nm wavelength at 20℃ . The PE effect of photo bleaching was the most significant under irradiating of laser among daylight lamp, light emitting diode（LED）, iodine-tungsten lamp and laser. For example, the photo bleaching rate of 240 μg/mL PE after 10 minutes’ irradiating under laser was close to the value using iodine-tungsten lamp of 240 minutes’ irradiating.

Key words: Porphyra yezoensis, Phycoerythrin, Purification, Molar extinction coefficient, Fluorescence quantum yield,Photobleaching

郭子叶,蔡春尔,李春霞,耿中雷,贾睿,何培民. 条斑紫菜高纯度藻红蛋白规模制备及光学性质研究[J]. 生物技术通报, 2013, 0(3): 192-198.

Guo Ziye, Cai Chuner, Li Chunxia, Geng Zhonglei, Jia Rui, He Peimin, . Pilot Preparation and Optical Properties of Highly Purified Phycoerythrin from Porphyra yezoensis[J]. Biotechnology Bulletin, 2013, 0(3): 192-198.

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条斑紫菜高纯度藻红蛋白规模制备及光学性质研究

Pilot Preparation and Optical Properties of Highly Purified Phycoerythrin from Porphyra yezoensis

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