Hepatoma-derived growth factor (HDGF) is a novel growth factor identified from conditioned medium of hepatoma cell line. HDGF expression is upregulated in many types of cancer. Besides, HDGF it is well known that stimulate the proliferation and migration of various types including fibroblasts, hepatoma cells, and endothelial cells. HDGF is composed of 240 amino acids with a well conserved, N-terminal PWWP (conserved Pro-Trp-Trp-Pro motif) domain (residues 1-100) and a highly variable C-terminal domain (residues 101-240). PWWP domain binds to heparin and heparin sulfate located outside the surface of cell membrane and facilitated internalization of the protein into cells. There are two putative bipartite nuclear localization signals (NLSs) in HDGF. The cell surface receptor of HDGF is not been identified so far. The amino acids 81 to 100 are responsible for NIH3T3 membrane receptor binding domain and Lys96 played a major role in this domain. In the present study, I expressed and purified a functional recombinant HDGF protein from E. coli. Recombinant HDGF protein stimulated the growth and migration of cervical cancer HeLa cells. Subsequently, site-directed mutagenesis was been completed on produce recombinant HDGF protein with mutations in both NLSs and receptor binding (K96A). Respectively, Deletion of NLS1 and NLS2 abolished the nucleus targeting of HDGF and abrogated the growth promoting as well as the chemotaxic capability of recombinant HDGF. Substitution of a single amino acid (K96A) within this peptide was sufficient to diminish it is binding to the cell surface and it is proliferated activity. In summary, both NLSs and K96 residue are important to biological functions of HDGF protein.
| Identifer | oai:union.ndltd.org:NSYSU/oai:NSYSU:etd-0820109-222240 |
| Date | 20 August 2009 |
| Creators | Chiu, Lin-Hui |
| Contributors | Jiin-Tsuey Cheng,, Ming-Hong Tai, Ching-Mei Hsu |
| Publisher | NSYSU |
| Source Sets | NSYSU Electronic Thesis and Dissertation Archive |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.lib.nsysu.edu.tw/ETD-db/ETD-search/view_etd?URN=etd-0820109-222240 |
| Rights | not_available, Copyright information available at source archive |
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