紫球藻藻红蛋白α、β亚基在毕赤酵母和大肠杆菌中的表达

Cloning and Expression of Phycoerythrin α and βSubunit Genes in Pichia pastoris and Escherichia coli

将紫球藻藻红蛋白α、β亚基基因(pe A、pe B)分别插入到毕赤酵母表达载体p AO815和大肠杆菌表达载体p ET-28a中.重组的共表达载体p AO815-pe A-pe B转化毕赤酵母(Pichia pastoris)GS115,经甲醇诱导、SDS-PAGE电泳及DOT-BLOT分析,发现藻红蛋白表达量低.p ET28a-pe A、p ET28a-pe B表达载体分别转入E.coli BL21,经IPTG诱导,Western Blotting分析,结果显示在约20.1 ku处有特异性蛋白条带,与融合蛋白(Pea-his、Peb-his)预期大小基本一致.

Phycoerythrin （PE） is one of the most important proteins involved in light capturing during photosynthesis in Porphyridium cruentum. Its potential biological activities had gained widespread attention. In this paper, the PE α and β genes were expressed of recombinant subunit gene peA and peB. Recombinant plasmids pAO815-peA-peB was transformed into Pichia pastoris GSll5 to make the recombinant strains GSl15-pAO815-peA-peB. PE-peA-peB co-expressing in GS115- pAO815-peA-peB was validated by SDS-PAGE and DOT-BLOT analysis. SDS-PAGE analy- sis could not indicate that the expression of PE protein in GSll5- pAO815-peA-peB. The result of DOT-BLOT indicated that the expression yield of PE protein was low. Recombinant plasmids pET28a-peA and pET28a-peB were transformed into E. coli BL21 to make two recombinant strains BL21-pET28a-peA and BI21-pET28a-peB, respectively. PE expressing in two recombinant strains was validated by SDS-PAGE and Western Blotting analysis. SDS-PAGE analysis indicated that the fusion protein （Pea-his, Peb-his） was mostly inclusion bodies. Specific expression of PE was confirmed by Western Blotting analysis.