文献类型：期刊文献

英文题名：Impact of oxygen supply on rtPA expression in Escherichia coli BL21 (DE3): Ammonia effects

作者：Wang, Hengwei[1]; Wang, Fengqing[1]; Wei, Dongzhi[1]

机构：[1] Newworld Institute of Biotechnology, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road P.O. Box 311, Shanghai 200237, China

年份：2009

卷号：82

期号：2

起止页码：249

外文期刊名：Applied Microbiology and Biotechnology

收录：EI(收录号：20090711905088)

语种：英文

外文关键词：Bottles - Amino acids - Nitrogen - Cell growth - Growth kinetics - Recombinant proteins - Ammonia - Batch cell culture - Ions - Oxygen supply

摘要：In shake flasks, good oxygen supply tended to decrease rtPA expression in media containing only yeast extract and tryptone, while oxygen limitation would increase rtPA synthesis in the same medium. Our data showed that though the drop of rtPA expression in the 20-ml cultures of LBG or 2YTG was accompanied with a severe acetate accumulation, it was actually caused by low ammonia. The rtPA expression level could be significantly improved by increasing culture ammonium ion up to 500 mM. The effects of exogenous high ammonia on cell growth and rtPA expression were further examined in shake flasks and a 4-l fermentor. The high ammonia had no significant impact on cell growth and oxygen respiratory activity but significantly depressed the activities of glutamine synthetase/glutamate synthase and glutamate dehydrogenase, suggesting that ammonium ion as a nitrogen source improved the protein expression by mediating ammonia-assimilating enzymes. We thus propose in our work that E. coli cells, which were grown to a certain density to produce rtPA, would undergo nitrogen starvation under the low ammonia conditions even when the organic nitrogen sources remained abundant. The scale-up of rtPA production from shake flasks to fermentors could be readily achieved in the media containing rich ammonium ion. ? 2008 Springer-Verlag.