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目的 :提高人免疫缺陷病毒 II型 ( HIV- 2 ROD) gag基因在毕赤酵母中的表达量。方法 :研究了转化子生长的培养条件 ,包括不同的甲醇诱导浓度、诱导时间、溶解氧、摇瓶转速及不同 p H值对人 HIV- 2 gag表达的影响。表达产物进一步通过盐析和 Sephadex G- 1 0 0分子筛层析柱层析分离纯化。结果 :最佳表达条件是 1 .0 %甲醇诱导 ,诱导时间 3d,大于 85 %的溶解氧 ,摇瓶转速2 5 0 r·min-1,p H6.0～ 6.5 ,目的蛋白表达量达到 2 1 mg· L-1。同时 ,经纯化后得到了纯度大于95 %的目的蛋白。结论 :获得了 HIV- 2 RODgag重组蛋白在巴斯德毕赤酵母系统中分泌表达的最佳表达条件。
Objective: To improve the expression of gag gene of human immunodeficiency virus type 2 (HIV-2 ROD) in Pichia pastoris. Methods: The culture conditions of transformants were studied, including the effects of different concentrations of methanol, induction time, dissolved oxygen, shake flask speed and different p H values on human HIV-2 gag expression. The expression product was further purified by salting-out and Sephadex G-100 molecular sieve chromatography. Results: The optimum conditions for the expression of the target protein were as follows: methanol 1.0%, induction time 3d, dissolved oxygen more than 85%, shaking speed 250 r · min-1, p H6.0 ~ 6.5, target protein expression reached 2 1 mg · L-1. At the same time, the target protein with the purity of more than 95% was obtained after purification. Conclusion: The optimal expression condition of HIV-2 RODgag recombinant protein secreted in Pichia pastoris system was obtained.