| Summary: | 碩士 === 國立清華大學 === 生物資訊與結構生物研究所 === 102 === Spirochetes of the genus leptospira often cause, with worldwide distribution, the serious zoonotic disease leptospirosis. There have been numerous advances in the understanding of the molecular pathogenesis of the infection the host, yet the mechanism of a viral factor of leptospira, LipL32, remains unclear. LipL32 is a lipoprotein and abundant on the outer membrane of leptospira. Leptospirosis occurs through the leptospira outer membrane proteins including LipL32 interact with the extracellular matrix (ECM). Fibronectin (FN) is a prominent component of the extracellular matrix molecules with which bacterial cell surface proteins interact. The binding of F30 to LipL32 has been observed by stains-all CD and enzyme-linked immunosorbent assay experiments. And Ca2+ can modulate fibronectin binding to LipL32. The heparin-binding site, fibronectin type III repeats 12-13 (FN1213), contains FN12 and FN13 in which takes a clump of positively charged residues. The structural analyses of LipL32 indicate that it possesses a large area of negative electricity. We have predicated the possible interaction of LipL32-FN1213 complex by a docking program. We provide several biochemistry methods such as Native-PAGE, chemical cross linking and ITC (Isothermal Titration Calorimetry) experiments to study the protein-protein interaction using LipL32 and FN1213 as an example. There may be indeed interaction between LipL32 and FN1213 by non-covalent hydrophobic interaction through the D-loop (142DDDDD146, D146/D149) of LipL32 and the positive region (R98/R99/R101/R115/K117/R146) of FN13. From this study we suggest the interaction between LipL32 and FN1213 should be week and non-specific hydrophilic interactions.
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