单 位：
浙江理工大学材料与纺织学院教育部先进纺织材料与制备技术重点实验室
关键词：
家蚕丝素;类丝蛋白;精氨酸-甘氨酸-天冬氨酸;盐析法
摘 要：
利用基因重组技术合成了具有良好细胞粘附活性的重组家蚕丝素-RGD融合蛋白质(Silk-RGD),探讨了其盐析的工艺条件。研究发现,在融合蛋白等电点pH8附近,当硫酸铵在溶液中的饱和度较低(10%)时,其蛋白质沉淀量随着温度的升高而增加;当硫酸铵的饱和度升至20%时,蛋白质沉淀量不再随着温度的变化而变化,但与低饱和度硫酸铵时相比,沉淀量明显增加;当硫酸铵的饱和度升至30%或以上时,沉淀中杂蛋白含量明显增多,不利于目的蛋白的纯化。Silk-RGD融合蛋白相对合理而且易操作的盐析纯化条件为:pH8,硫酸铵在溶液中的饱和度为20%,室温(约20℃)。利用该工艺条件,可以低成本、高效率地生产Silk-RGD融合蛋白,从而满足新型材料规模化开发的需要。
译 名：
Analysis on Salting-out Purification of the Bombyx mori Silk Fibroin-RGD Fusion Protein
作 者：
LI Yuan BAI Li-Qiang YAO Ju-Ming*(The Key Laboratory of Advanced Textile Materials and Manufacturing Technology of Ministryof Education,Collage of Materials and Textile,Zhejiang Sci-Tech University,Hangzhou 310018,China)
关键词：
Bombyx mori silk fibroin;Silk-like protein;RGD;Salting-out
摘 要：
By applying of genetic engineering strategies a Bombyx mori silk fibroin-RGD fusion protein(Silk-RGD) was synthesize,which showed a high cell adhesive ability,and the technological conditions for salting-out purification of the Silk-RGD were studied.The results revealed that the yield of salting-out precipitate increased with rise of temperature at the low(NH4)2SO4 concentration(10%) under pH 8 of isoelectric point;when the(NH4)2SO4 concentration increased to 20%,the yield was not temperature-dependent but much more than that at concentration of 10%;when the(NH4)2SO4 concentration increased to 30% or above,there were many impurities in the precipitates causing difficulties for purification of the target protein.As a conclusion,it seemed that the rational and easy operating salting-out conditions for purification of Silk-RGD were 20% of(NH4)2SO4 at room temperature(around 20 ℃) and pH 8.Under these conditions,Silk-RGD can be produced with high efficiency and low cost to meet the demands of development of new material in large scale.
