摘要
由重组大肠杆菌表达的人粒细胞 -巨噬细胞集落刺激因子 (rh GM- CSF)通过包涵体抽提、凝胶过滤、复性、疏水和离子交换层析获得纯度达到 97%的产品 ,比活为 3.2× 10 7IU/ mg,分子量为 14.84 k D.纯化获得的 rh GM- CSF为一酸性蛋白 ,等电点约为 5.2 ,NH2 ,-末端序列测定与文献报道一致 ,NH2 -末端不均一 ,其中带 Met的分子约占 59.5% ,成熟蛋白分子约占 2 5%。蛋白的纯化倍数为 3.9,生物活性回收率为 85.3%。
Human granulocyte macrophage colony stimulating factor expressed by recombinant E. coli cells was purified to homogeneity by extraction and solubilization of inclusion bodies, gel filtration,renaturation,hydrophobic interaction and ion exchange chromatographic procedure.The purified rhGM CSF has attained the criteria of purity level about 97%,has a specific activity of about 3.2×10 7 units/mg,molecular weight is about 14.84kD.rhGM CSF is an acidic protein (pI5.2),partial NH 2 terminal sequence are ideatical with those reported for this protein. The results also showed that it is apparently heretogeneous from its NH 2 terminal side as it is composed of two polypeptides having Met and Ala as the NH 2 terminal residues in which the intact Met analogue accounts for 59.5%,Ala analogue accounts for 25%. The overall purification factor obtained was about 3.9,the recovery of purified rhGM CSF was 83.3% by a biological assay method.
出处
《药物生物技术》
CAS
CSCD
1997年第2期72-76,共5页
Pharmaceutical Biotechnology
关键词
粒细胞
巨噬细胞
集落刺激因子
纯化
鉴定
Granulocyte macrophage colony stimulating factor,Purification, Characterization
