文献类型：期刊文献

中文题名：诱导相温度对毕赤酵母表达重组人复合α干扰素聚合的影响

英文题名：The Effect of Induction Temperature on Aggregation of Consensus Interferon-α Expressed by Pichia pastoris

作者：史琪琪[1];郝玉有[1];吴康华[2];储炬[1];庄英萍[1];张嗣良[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237;[2]上海贸基生物技术有限公司,上海200336

年份：2006

卷号：22

期号：2

起止页码：311

中文期刊名：生物工程学报

外文期刊名：Chinese Journal of Biotechnology

收录：CSTPCD;;Scopus;北大核心:【北大核心2004】；CSCD:【CSCD2011_2012】；PubMed;

基金：国家高技术研究发展计划("863计划"项目)(No.2002AA217021;2002AA2Z3451)资助。~~

语种：中文

中文关键词：诱导相温度;重组人复αa干扰素;聚合体;毕赤酵母

外文关键词：induction temperature, consensus interferon α, aggregation, Pichia pastoris

摘要：研究了Pichia pastoris表达重组人复合α干扰素(cIFN)时诱导期温度对cIFN形成聚合体的影响。在5L罐上考察毕赤酵母在不同温度(30、25、20℃)下诱导时菌体生长和cIFN表达的差异,发现毕赤酵母在20℃下菌体生长不受影响,总蛋白表达量有所降低,相当于30℃发酵时胞外总蛋白的67.8%。但是通过非还原性电泳、Native-PAGE及Western blotting分析发现较高温度(30℃)诱导表达时,分泌到胞外的cIFN容易形成聚合体,单体含量很少。诱导相控制温度为20℃时,明显降低了cIFN聚合体的形成,cIFN单体量为570mg/L,发酵上清液抗病毒活性为1.05×109IU/mL。与控制诱导相温度为30℃相比,cIFN单体量提高了7.2倍,发酵上清液中单位体积的cIFN抗病毒活性提高了38.7倍。
The effect of induction temperature on aggregation of consensus interferon-α expressed by Pichia pastoris was investigated. The cell growth and cIFN level were analyzed and compared when Pichia pastoris was grown at 30,25,20℃ during induction phase, using 5.0L fermentor, The result suggested that the cell growth was not affected much under the different induction temperature, but the protein level declined markedly with the decrease of the induction temperature. The total protein ammount induced at 20℃ was 67.8 percent of that at 30℃. SDS-PAGE and native-PAGE as well as Western blotting analysis were further conducted. The electrophoresis results revealed that cIFN formed aggregates after secreted into media when protein was induced at 30℃, but this problem can be restored by decreasing the induction temperature to 20℃. cIFN monomer in supernatant arrived at 570mg/L and bioactivity of fermentation broth reached 1.05×10^9IU/mL at 20℃ of induction temperature. The amount of clFN monomer and bioactivity in supernatant elevated 7.2 and 38.7 times, respectively, when the induction temperature was controlled at 20℃ instead of conventional 30℃.