23 バクテリア複合糖質リピドAの標識化類縁体の合成と生物機能(口頭発表の部)

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To elucidate the mechanism of the biological events caused by bacterial endotoxin (lipopolysac-charide, LPS), isotope and fluorescence labelings were effected on lipid A, which is the partial structure of LPS essential for its bioactivity. The labelings were performed by means of chemical synthesis, and physicochemical as well as biological functions of the labeled derivatives were evaluated. 6-^<13>C-Labeled derivatives were synthesized of a biosynthetic precursor of lipid A (6-^<13>C-2) and its analogue with shorter (C10) acyl chains (6-^<13>C-3). NMR study of the biosynthetic precursor 2 using ^<13>C-labeled and unlabeled specimens enabled us to figure out its supramolecule structure formed by self-assembly. By contrast, the short-acyl analogue 3 was found to form no aggregate. Molecular modeling revealed the origin of this striking behavioral and conformational difference: unnatural C10 length for the acyl moieties is not sufficient to obtain stabilization by hydrophobic interaction. Because it has already shown that the short-acyl analogue 3 no longer retains the biological activity of the biosynthetic precursor 2, it is strongly suggested such a supramolecule-forming ability relates with the biological activity of lipid A analogues. The fluorescence-labeled lipid A analogue 5 was next synthesized to obtain a deeper insight on the self-assembly of lipid A. BODIPY^[○!R] was employed as a fluorescence group, and the labeling was effected on the phosphonooxyethyl (PE) analogue 4 wherein the chemically labile glycosyl phosphate of lipid A is replaced with the stable 2-(phosphonooxy)ethyl group. The synthesis was achieved using allyl-type groups for the persistent protection of all active functionalities to afford the highly pure final product. The labeled analogue 5 was found to retain definite activities of lipid A1. Fluorescence spectra indicated the critical micelle concentration of 5 being below 0.46nM. From these experiments it can be clearly concluded that the labeled lipid A analogue 5 forms aggregate around the concentration range where it exhibits the biological activities. To identify the possible receptor(s) on the competent animal cells, tritium-labeling was effected on the ethylene glycol moiety of the PE analogue 4 of lipid A. Careful two-step oxidation of the ally group and partition chromatographic purification after final deprotection led to complete removal of over-oxidized by-products and eventually enabled us to obtain a highly pure (>98% radiochemical purity) preparation of ^3H-4 with high specific radio activity (62GBq/mmol). Using the ^3H-labeled and unlabeled PE analogues, four binding proteins were detected in macrophages from both LPS-responder C3H/HeN mice and LPS-hyporesponder C3H/HeJ mice.
天然有機化合物討論会の論文
1999-09-01

23 バクテリア複合糖質リピドAの標識化類縁体の合成と生物機能(口頭発表の部)

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