The focus of this research is to describe the production and characterization of the human granulocyte-macrophage colony-stimulating factor (hGM-CSF) in insect cells, using Autographa californica buclear polyhedrosis virus (AcNPV) as an expression vector. All three forms of biological activity of hGM-CSF. Following N-glycanase treatment, the two glycosylated hGM-CSF proteins (15.5 and 16.5 KDa) which bound to Concanavalin A affinity column ran as a 14.5-15.5 KDa band on SDS-PAGE. Western blot analysis of expression in Sf9 cells treated with tunicamycin revealed only the presence of the 14.5 KDa species. The N-terminal amino acid sequence of the recombinant hGM-CSF was identical to that of natural hGM-CSF deduced from cDNA. These results demonstrate that baculovirus-produced hGM-CSF could be N-glycosylated in Sf9 cells, the signal peptide of recombinant hGM-CSF could be recognized and cleaved by infected insect cells and the resultant molecule secreted into the medium.
| Identifer | oai:union.ndltd.org:unt.edu/info:ark/67531/metadc798471 |
| Date | 12 1900 |
| Creators | Chiou, Chuang-Jiun |
| Contributors | Wu, Edward Ming-chi, 1938-, Benjamin, Robert C., Pirtle, Robert M., O'Donovan, Gerard A., Knesek, John, Goven, Arthur James, 1950- |
| Publisher | University of North Texas |
| Source Sets | University of North Texas |
| Language | English |
| Detected Language | English |
| Type | Thesis or Dissertation |
| Format | x, 112 leaves : ill., Text |
| Rights | Public, Chiou, Chuang-Jiun, Copyright, Copyright is held by the author, unless otherwise noted. All rights |
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