Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Expression of Murine GM-CSF in Recombinant Aspergillus niger

  • Kim, Nyoung-Ji (Institute for Molecular Biology and Genetics, Chonbuk National University) ;
  • Kwon, Tae-Ho (Institute for Molecular Biology and Genetics, Chonbuk National University) ;
  • Jang, Yong-Suk (Institute for Molecular Biology and Genetics, Chonbuk National University) ;
  • Yang, Moon-Sik (Institute for Molecular Biology and Genetics, Chonbuk National University) ;
  • Kim, Dae-Hyuk (Institute for Molecular Biology and Genetics, Chonbuk National University)
  • Published : 2000.06.01
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Abstract

Recombinant Aspergillus niger was constructed to express and secrete a biologically active murine granulaocyte macrophage-colony stimulating factor (mGM-CSF). A 500 bp fragment encoding the signal peptide and terminator of glyceraldehyde-3-phosphate dehydrogenase (gpd). The hygromycin phosphotrasferase gene (hph) was used as a selection marker for the fungal transformants. An expression vector was introduced into A. niger ATCC 9642, and a Northern blot analysis indicated the presence of a considerable amount of transcripts from the introduced mGM-CSF. The biological activity of recombinant mGM-CSF (rmGM-CSF) isolated from the culture filtrate was confirmend by measuring the proliferationof the GM-CSF dependent FDC-P1 cell line. It appeared that rmGM-CSF was amenable to the proteolytic activity produced by A. niger, since biological actibity was only observed when the transformants were grown in a protease-repressing medium, and the activity of rmGM-CSF dramatically decreased with an increase of age of the culture. The yield of rmGM-CSF, as determined by ELISA. was 640 ng/l of culture filtrate. Accordingly, its specific activity is estimated to be approximately two-and-a-half times higher than that of a commercial preparation from E. coli.

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