89 プロスタサイクリン受容体タンパク質の構造研究 : 光親和性標識法による分子量の同定(口頭発表の部)

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The azidophenyl derivative, (15S)-10c. has been synthesized by structural modification of isocarbacyclin methyl ester (3). First, the C(13)-C(14) double bond of 3 is selectively epoxidized by Sharpless epoxidation giving 4, whose 11- and 15-hydroxyl groups are acetylated. Epoxy ring opening of resulting 5 with CH_3COOH-H_2O and subsequent deacetylation with aqueous K_2CO_3 give 6, which is cleaved with NaIO_4 to give an aldehyde 7. Horner-Emmons reaction with 8c gives an enone 9c. Reduction of 9c with NaBH_4-CeCl_3 is followed by separation of the resulting 15-epimers and alkaline hydrolysis of the methyl ester to give (15S)-10c. This compound exhibits high affinity to the PGI_2 receptor protein(s) in mastocytoma P-815 cells with the IC_<50> value of 3nM. The tritium labeled derivative, [^3H]-(15S)-10c, synthesized by reduction of 9c with [^3H]NaBH_4-CeCl_3 followed by alkaline hydrolysis, has been used for the photoaffinity labeling experiment. Plasma membrane fraction of mastocytoma P-815 cells which is abundant in the PGI_2 receptor protein(s) is incubated with [^3H]-(15S)-10c and then irradiated with a UV lamp. The irradiated material is subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and fluorography showing a clear band around 43k. The photoreaction in presence of GTPγS decreases the intensity of this band. The addition of iloprost (11) to the incubated media completely suppresses the formation of this band. These results confirm the molecular weight of the PGI_2 receptor protein(s) in mastocytoma P-815 cells to be 43k.
1991-09-07

89 プロスタサイクリン受容体タンパク質の構造研究 : 光親和性標識法による分子量の同定(口頭発表の部)

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