Improved process for production of recombinant yeast-derived monomeric human G-CSF |
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Authors: | C S Bae D S Yang J Lee Y-H Park |
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Institution: | (1) Biochemical Process Engineering R.U., Korea Research Institute of Bioscience and Biotechnology (KRIBB), P.O. Box 115, Yusong, Taejon 305-600, South Korea, KR;(2) Bioprocess Engineering Laboratory, Hanhyo Institutes of Technology, Taejon 305-309, South Korea e-mail: jwlee@kribb4680.kribb.re.kr Tel.: +82-42-860-4449 Fax: +82-42-860-4594, KR |
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Abstract: | The human granulocyte colony-stimulating factor (hG-CSF) was efficiently secreted at high levels in fed-batch cultures of
recombinant Saccharomyces cerevisiae. However, the secreted recombinant hG-CSF (rhG-CSF) was shown to exist as large multimers in the culture broth due to strong
hydrophobic interaction. It was hardly monomerized even by urea at high concentration. This multimer has been reported to
diminish specific receptor-binding activity of hG-CSF and causes undesirable problems in the downstream process. When the
rhG-CSF was secreted to extracellular broth in the presence of a non-ionic surfactant (Tween 80) in the culture media, the
multimerization of the secreted rhG-CSF was efficiently prevented in the fed-batch cultures. Also, the monomer fraction and
secretion efficiency of rhG-CSF were significantly increased at the higher culture pH (6.5). Without using any denaturing
agents, the secreted rhG-CSF monomer was easily purified with high recovery yield and purity via a simple purification process
under acidic conditions, consisting of diafiltration, cation exchange, and gel filtration chromatography. A lyophilization
process devoid of intermonomer aggregation was also designed using effective stabilizing agents.
Received: 2 March 1999 / Received revision: 16 April 1999 / Accepted: 23 April 1999 |
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