雷竹SOC1蛋白原核表达及纯化

Expression and purification of Phyllostachys violascens SOC1 protein in prokaryotic system

为了获得有活性的雷竹Pv SOC1蛋白（Phyllostachys violascens SUPPERSSOR OF CO OVEREXPRESSION 1）,并进一步讨论其结构形态,通过原核表达方法得到Pv SOC1的可溶性重组蛋白,以p ET-GST作为表达载体,大肠杆菌E.coli Rosetta作为宿主菌株,超表达全长和IKC结构域的雷竹Pv SOC1蛋白。发现在37℃条件下,菌液浓度D（600）值达到0.4-0.6时进行诱导,控制IPTG终浓度为0.4 mmol/L,诱导目的蛋白表达5 h,可以获得可溶的IKC结构域GST融合蛋白。采用TEV（tobacco etch virus protease）酶切技术、配合GST亲和层析柱及分子层析色谱柱,去除标签蛋白并纯化目的蛋白。所获得的高纯度IKC结构域Pv SOC1蛋白经过对比分析发现其以八聚体形式存在。

In order to obtain soluble recombinant Pv SOC1（ Phyllostachys violascens SUPPERSSOR OF CO OVEREXPRESSION 1） protein in vitro. We used p ET-GST as expression vector,to over-express full-length and IKC structural domain of Pv SOC1 in E.coli Rosetta prokaryotic system. We found the optimal expression strategy is when the suspension culture＇s OD600 value reaches 0.4-0.6,add IPTG and keep it＇s concentration at 0.4 mmol / L. Kept incubating for 5 h at the temperature of 37℃ to induce the protein.We found IKC domain is soluble,to compare with the fullength sequence,it started from No.62 amino acid and ends at No. 225. Added TEV（ tobacco etch virus protease） protease to remove GST tag.Combined glutathione sepharose resin with size exclusion chromatography（ SEC） to get highly purified IKC domain of Pv SOC1.And chromatography data proof this domain exists as a octamer.