鹰嘴豆α-淀粉酶抑制剂基因CL-AI的原核表达、纯化及生物学特性

发布时间：2018-04-11 04:22

本文选题：α-淀粉酶抑制剂 + 原核表达　；参考：《核农学报》2017年05期

【摘要】：为研究鹰嘴豆种子中新贮藏蛋白类型α-淀粉酶抑制剂CL-AI的抑制活性、生物活性以及三级结构,本研究利用前期克隆的α-淀粉酶抑制剂基因CL-AI及其2个亚基CL-AI-α、CL-AI-β分别构建原核表达载体p E-SUMO3,经诱导表达获得了CL-AI、CL-AI-α的可溶性融合蛋白和CL-AI-β包涵体融合蛋白,其中CL-AI-β通过构建含不同标签的表达载体,最终实现了可溶性表达。人唾液淀粉酶(HSA)抑制活性试验结果表明,CL-AI,CL-AI-α,CL-AI-β3个蛋白对HAS都有一定的抑制作用。发芽试验表明,CL-AI蛋白对小麦、玉米、水稻种子发芽过程中的发芽势、根长和芽长影响显著,与加入无菌水的处理相比,种子的简化活力指数显著下降。本研究结果为CL-AI蛋白的后续研究与应用奠定了基础。
[Abstract]:In order to study the inhibitory activity, biological activity and tertiary structure of 伪 -amylase inhibitor CL-AI, a new type of storage protein in chickpea seeds,In this study, the prokaryotic expression vector pE-SUMO3 was constructed by using the previously cloned 伪 -amylase inhibitor gene CL-AI and its two subunits, CL-AI- 伪 and CL-AI- 尾, respectively. The soluble fusion protein and CL-AI- 尾 inclusion body fusion protein of CL-AICL-AI- 伪 and CL-AI- 尾 inclusion body fusion protein were obtained by induction expression.CL-AI- 尾 was finally expressed in soluble form by constructing expression vector containing different tags.Human salivary amylase (HSA) inhibitory activity test showed that CL-AI- 伪 CL-AI- 尾 3 proteins could inhibit HAS to some extent.Germination test showed that CL-AI protein had a significant effect on germination potential, root length and bud length of wheat, maize and rice seeds during germination. Compared with the treatment of aseptic water, the simplified vigor index of seeds decreased significantly.The results of this study laid a foundation for the further study and application of CL-AI protein.
【作者单位】：南京农业大学作物遗传与种质创新国家重点实验室;新疆农业大学干旱区荒漠研究所;新疆农业大学农学院;
【基金】：国家自然科学基金(30960206;31160306)
【分类号】：S529

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