Method for large-scale isolation and purification of R-phycoerythrin from red alga Polysiphonia urceolata Grev.
R-phycoerythrin was isolated and purified from a red alga, Polysiphonia urceolata Grev, using Streamline column combined with ion-exchange chromatography or hydroxyapatite chromatography. The purity of R-phycoerythrin isolated by Streamline column was up to 1.66 and the yield of R-phycoerythrin could be as high as 0.68 mg/g frozen P. urceolata. All the eluates from Streamline column were divided into two equivalent parts, respectively. One part was pumped into the ion-exchange column loaded with Q-Sepharose and the other was applied to the adsorption column loaded with hydroxyapatite. The purities of R-phycoerythrin purified using these two methods were both up to 3.26, more than 3.2 the commonly accepted criterion. The yield of purified R-phycoerythrin from the ion-exchange chromatography was 0.40 mg/g frozen P. urceolata and that from the hydroxyapatite chromatography could reach 0.34 mg/g frozen P. urceolata. The purified protein had three absorption peaks at 498, 535, and 565 nm and displayed a fluorescence maximum at 580 nm, which was consistent with the typical spectrum of R-phycoerythrin. The purified R-PE was also identified with electrophoresis. Only one single protein band appeared on native-PAGE with silver staining. SDS-PAGE demonstrated the presence of one 20 kDa major subunit, and one low intensity band corresponding to 33 kDa subunit. The results indicate that using the expanded bed adsorption combined with ion-exchange chromatography or hydroxyapatite chromatography, R-phycoerythrin can be purified from frozen P. urceolata on large scale.
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Nanhai Road 7, Qingdao 266071, China.
Jian-Feng
JF
Niu
Guang-Ce
GC
Wang
Cheng-Kui
CK
Tseng
D000645:Ammonium Sulfate; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D010799:Phycoerythrin / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D000461:Rhodophyta / Q000235:genetics / Q000378:metabolism*; D012965:Sodium Chloride; D013057:Spectrum Analysis
D010799:Phycoerythrin; D012965:Sodium Chloride; D000645:Ammonium Sulfate