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目的获取纯化的霍乱弧菌HutX蛋白晶体和衍射数据。方法通过基因克隆和蛋白表达得到HutX蛋白,通过镍柱亲和层析、Source Q阴离子交换柱层析和Superdex 200分子筛柱层析等纯化所得蛋白,并用蛋白质印迹法进行验证。将所获蛋白进行结晶条件筛选及悬滴优化,X线衍射分析所得晶体的结构。结果蛋白质印迹分析间接证明所得到的蛋白为HutX蛋白,并得到HutX蛋白的晶体及衍射数据。结论为进一步研究HutX的蛋白结构及其在霍乱弧菌亚铁血红素利用机制中发挥的作用提供了参考。
Objective To obtain purified crystals and diffraction data of HutX protein from V. cholerae. Methods HutX protein was obtained by gene cloning and protein expression. The purified protein was purified by nickel affinity chromatography, Source Q anion exchange chromatography and Superdex 200 column chromatography. The protein was identified by Western blotting. The obtained protein was subjected to crystallization conditions selection and pendant drop optimization, and the resulting crystal was analyzed by X-ray diffraction. Results Western blot analysis indirectly demonstrated that the obtained protein was HutX protein and obtained the crystal and diffraction data of HutX protein. Conclusion This study provides a reference for further study on the protein structure of HutX and its role in the mechanism of utilization of heme in Vibrio cholerae.