| Autor: |
Vella, F., Hernandez, J.F., Arlaud, G.J., Molla, A., Block, M.R. |
| Předmět: |
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| Zdroj: |
Journal of Peptide Research; Nov99, Vol. 54 Issue 5, p415-426, 12p |
| Abstrakt: |
Abstract: A novel protein was engineered by inserting the GRGDS motif of fibronectin within the 14-residue loop of the EGF-like module from human complement protease C1r. The resulting chimeric EGF-RGD module (52 residues, three disulfide bridges) was assembled by automated solid-phase synthesis using the t-Boc strategy. Using reduced/oxidized glutathione, the EGF-RGD module was folded as efficiently as the natural C1r-EGF module, resulting in formation of the appropriate disulfide bridge pattern as shown by mass spectrometry and N-terminal sequence analyses of thermolytic fragments. Circular dichroism and NMR measurements provided further indication that introduction of the GRGDS motif had no significant effect on the folding. Using Chinese Hamster Ovary (CHO) cells bearing the integrin receptors specific for fibronectin and vitronectin, EGF-RGD was shown to induce cell adhesion via the introduced GRGDS motif. Cell binding was inhibited specifically and efficiently by the synthetic peptide GRGDSP and by fibronectin, and to a much lesser extent by vitronectin, whereas the monoclonal antibody PB1 directed to the α5 subunit of α5β1 integrin had no effect. The ability of EGF-RGD to trigger significant cell spreading and intracellular signaling was also demonstrated using immunofluorescence and confocal microscopy. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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