发菜藻蓝蛋白分离纯化的研究

以发菜为材料,比较了提取液类型和饱和硫酸铵浓度对藻蓝蛋白提取的影响,并对藻蓝蛋白的提取程序和部分特性进行了研究。结果表明:50 mmol/L KP缓冲液(pH值7.2)是合适的提取液,体积分数为40%~50%饱和硫酸铵盐析效果优于其它浓度。经过DEAE-Toyopeal 650 S离子交换层析和SuperdexTM200凝胶过滤层析后,藻蓝蛋白纯度达6.2,最大吸收峰位于615 nm,荧光发射峰位于649 nm,由α和β2个亚基组成,其分子质量分别为18 051.17和19 142.27 Da。因此,发菜藻蓝蛋白分离纯化较为理想的程序为:藻粉→50 mmol/L KP缓冲液(pH值7.2)浸泡→French pressure(1 500 kg/cm2)破碎细胞→40%~50%饱和硫酸铵盐析→DEAE-Toyopeal 650 S离子交换层析→SuperdexTM200凝胶过滤层析→较纯的藻蓝蛋白。

Isolation and Purification of C-phycocyanin from Nostoc flagelliforme

Nostoc flagelli forme is a kind of terrestrial cyanobacterium of great economic value.C-phycocyanin is a kind of blue protein belonging to the photosynthetic apparatus,and of fluorescence molecular probe both with the characteristics of absorption spectra and fluorescence spectra.In the study,with N.Flagelliforme as materials,the effects of the types of extraction liquid and concentration of ammonium sulfate on　the extraction of C-phycocyanin were compared.An efficient procedure for isolation and purification of C-phycocyanin was studied and some of its properties were also investigated.The results showed that 50 mmol/L K phosphate buffer(pH 7.2)was more ideal extraction liquid.With solid ammonium sulfate at 40%～50%saturation was more ideal concentration for precipitation,and then followed by ion exchange chromatography on DEAE-Toyopeal 650 S column and gel filtration chromatography on SuperdexTM 200 column,pure C-phycocyanin was finally obtained from N.Flagelliforme with purity ratio(A615/A280)6.2.The max absorption and fluorescence emission of purified C-phycocyanin was at 615 am and at 649 nm re-spectively.C-phycocyanin was composed of α and β subunits,and the molecular mass of α and β subunits were 18 051.17 and 19 142.27 Da respectively.Therefore,the more ideal procedure of extracting C-phycocyanin from N.Flagelliforme was as follows:algae power→soaking with 50 mmol/L K phosphate buffer(pH 7.02)→crushing cells by French pressure(1 500 kg/cm2)→salting out with 40%～50%saturated ammonium sulfate→DEAE-Toyopeal 650 S column chromatography→SuperdexTM 200 column chromatography→gaining C-phycocyanin with high purity.