纯化标签不同融合端的选择对蝎毒肽色谱行为的影响

目的考察组氨酸标签置于抗癫痫肽AEP肽链不同末端对重组肽亲合层析的影响。方法同源建模构建AEP三维结构,预测空间构象对纯化标签的屏蔽效应;构建组氨酸标签不同末端融合表达载体;对比两者相同色谱条件下的层析行为及生物活性。结果 AEP构象分析显示,肽链C末端空间屏蔽效应小;在两者表达量相当的前提下,经亲和层析纯化,每升发酵液可获色谱纯样品,AEP-His6为3.3mg,Met-Gly-His6-AEP为0.8mg;HistagC端融合对rAEP活性影响较N端融合小。结论对于蝎毒这类空间结构简单的小分子活性肽,组氨酸标签融合端的选择对重组肽色谱行为及活性有显著影响。

The effect of the location of purification tag fusion on chromatography behavior and activity of recombinant scorpion venom peptides

Objective To investigate the chromatography behavior of N- or C-terminal His tag fusion recombinant anti-epilepsy peptide （AEP）. Methods Three dimensional model of AEP was built by homology-based method. The shielding effect of His tag by spatial conformation was predicted, and fusion peptides were successfully expressed and purified. Biological activity of them were assayed. Results A conformational analysis of AEP showed that the space shielding effect of the C-terminal region of fusion tag was less potent than that of the N-terminus. Under the premise of the same expression amount and chromatographic conditions of N- or C-terminal His tag fusion recombinant AEP, the production of AEP-His6 and Met-Gly-His6-AEP was 3.3 mg and 0.8 mg per liter fermentation broth, respectively. The effect on rAEP activity by the His tag C-terminal fusions was less significant than that of the N-terminal fusion. Conclusions The place of His tag at N- or C-terminus has significant impact on chromatographic behavior and activity of small molecule active peptides like scorpion venom which possesses simple spatial conformation.