Abstract:

Under a suitable condition of crystallization, dark brown short rhombohedron crystals could be obtained from nitrogenase MnFe protein purified from a mutant UW3 of Azotobacter vinelandii Lipmann grown in Mn-containing but Mo- and NH3-free medium. The possibility of crystallization, and number,size and quality of crystals were obviously dependent on concentrations of NaCl,MgCl2, PEG 8000,Tris and Hepes buffer and on methods for crystallization. PEG concentration affected on the shape of the crystals.The optimal concentrations of the chemicals for crystallization of MnFe protein were slightly different from those for crystallization of ΔnifZ MoFe protein from a nifZ deleted strain of Azotobacter vinelandii .　SDS-PAGE showed that the protein from the dissolved crystals was almost the same as MnFe protein before crystallization, indicating that the crystal was formed from MnFe protein.

固氮酶锰铁蛋白的晶体生长
黄巨富¹董志刚¹汪道涌¹吕玉兵¹ 张华峰¹王耀萍²韩毅² 代小虎²

（1. 中国科学院植物研究所，北京100093；2.  中国科学院生物物理研究所，北京100101）

摘要：从以Mn代钼的固氮培养基中固氮生长的固氮菌(Azotobacter vinelandii Lipmann)突变种UW3中分离纯化的MnFe蛋白，在一定的结晶条件下，可从溶液中析出深棕色的短斜四棱柱晶体。Tris和Hepes缓冲液、NaCl、MgCl2和PEG的浓度及结晶方法等，对该蛋白的出晶率、晶核数目、晶体大小和质量均有明显的影响，PEG浓度的改变还可使该蛋白晶体的晶型发生变化。MnFe蛋白结晶所需的上述化合物的最适浓度与缺失nifZ固氮菌突变种ΔnifZ MoFe蛋白结晶所需的最适浓度有所不同。SDS凝胶电泳表明，晶体溶解后的蛋白与结晶前的MnFe蛋白基本相同。结果表明，该晶体为MnFe蛋白的晶体。

关键词： 固氮菌突变种UW₃；含锰培养基；固氮酶锰铁蛋白；晶体生长

Key words: mutant UW3 of Azotobacter vinelandii, Mn-containing medium, nitrogenase MnFe protein, crystalline growth