%0 Journal Article %T Preparation and identification of recombinant human interferon-γ
重组人干扰素-γ的制备与鉴定 %A WU Dan %A GAO Dong %A BAI Quan %A GENG Xindu %A
吴丹 %A 高栋 %A 白泉 %A 耿信笃 %J 色谱 %D 2008 %I %X The renaturation with simultaneous purification of recombinant human interferon-gamma (rhIFN-gamma) expressed as inclusion bodies in Escherichia coli (E. coli) was accomplished by the stationary phase of hydrophobic interaction chromatography (STHIC) with the end group of poly(ethylene glycol) (PEG)(PEG200) packed in a chromatographic column and a chromatographic pie by nonlinear gradient, separately. In order to provide more selections for the chromatographic separation of rhIFN-gamma from different sources, the chromatographic behavior of rhIFN-gamma in reversed-phase liquid chromatography, ion-exchange chromatography and immobilized-nickel affinity chromatography were also studied. The fraction of the renatured and purified rhIFN-gamma from HIC was desalted by the size exclusion chromatography, subsequently freeze-dried to powder. With matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS), monomeric and dimeric rhIFN-gamma were found in the powder due to the freeze-dried process and their relative molecular masses were 17 184.0 and 34 204.4, respectively. With the bioactivity assay by cytopathic effect inhibition (CPEI), the specific bioactivity of rhIFN-gamma was 9.5 x 10(8) IU/mg, which was higher than that of the required criteria in the phar macopoeia of China, because the presence of dimeric rhIFN-gamma which has much higher specific bioactivity than its monomer in the powder. The obtained mass recovery, purity, specific bioactivity of the purified monomeric rhIFN-y were 93.7%, > 95%, and 4.3 x 10(7) IU/mg, respectively. The results showed that the renaturation with simultaneous purification of rhIFN-gamma by PEG200-STHIC is a kind of efficient method. %K 重组人干扰素-γ %K 蛋白折叠液相色谱法 %K 高效疏水色谱 %K 复性 %K 纯化 %K 重组人干扰素 %K 制备与鉴定 %K human %K recombinant %K identification %K 方法 %K 结果 %K 质量回收率 %K 柱复性 %K 凝胶电泳 %K 聚丙烯酰胺 %K 十二烷基硫酸钠 %K 比活性 %K 抑制法 %K 细胞 %K 二聚体 %K 相对分子质量为 %K 单体 %K 测定 %K 电离飞行时间质谱 %U http://www.alljournals.cn/get_abstract_url.aspx?pcid=6E709DC38FA1D09A4B578DD0906875B5B44D4D294832BB8E&cid=6579068328FE643F&jid=4D81E042D77AFEC6881D14759692069C&aid=05117C2081B09A4F2ACF5BCD547669A7&yid=67289AFF6305E306&vid=96C778EE049EE47D&iid=0B39A22176CE99FB&sid=9F6DA927E843CD50&eid=CC0ECB9C52F1B85F&journal_id=1000-8713&journal_name=色谱&referenced_num=0&reference_num=23