| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 2002: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 2001: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Research Abstract |
The synthetic lipopeptide FSL-1 [S-(2,3-bispalmitoyioxypropyl)-Cys-Gly-Asp-Pro-Lys-His-Pro-Lys-Ser-Phe, Pam_2 CGDPKHPKSF] synthesized on the basis of the structure of the N-terminal lipopeptide moiety of a Mycoplasma salivarium lipoprotein activated human gingival fibroblasts to induce the cell surface expression of ICAM-1 and the production of IL-6, IL-8 and monocyte chemoattractant protein-1. FSL-1 also activated THP-1 cells, a monocytic cell line, to produce TNF-α. Both the diacylglyceryl cysteine (Pam_2C) and peptide (CGDPKHPKSF) portions of FSL-1 failed to activate THP-1 cells. A single amino acid substitution from Phe to Arg at the C terminus of FSL-1 and a fatty acid substitution from pralmitic acid to stearic acid drastically reduced the activity. The data obtained by experiments using Chinese hamster ovary cells transfected with CD14, CD14/Toll-like receptor 2 or CD14/Toll-like receptor 4 genes suggested that signaling by FSL-1 was mediated by Toll-like receptor 2. Mycoplasmal diacylated lipopeptides, FSL-1 and MALP-2 (Pam_2CGNNDESNISFKEK) and bacterial triacylated lipopeptides, S-(2 ,3-bispalmitoyloxypropyl)-N-palmitoyl-Cys-Ser-Asn-Asn-Ala (Pam_3 CSNNA) and Pam_3CSK_4 were examined for their abilities to activate THP-1 cells. FSL-1, MALP-2, Pam_3CSK_4 and Pam_3CSNNA had the activitiy to induce TNF-α Production by THP-1 cells, but the level of the activity decreased in this order.
These results suggest that both the diacylglyceryl and peptide portions of FSL-1 are essential for the expression of biological activity, that the differences in fatty acid in the diacylglycerol portion and amino acid sequence of the peptide portion affect the biological activity, and that the electrostatic interaction plays an important role in the recognition of microbial lipopeptides by Toll-like receptor 2.
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