TAILIEUCHUNG - Báo cáo khoa học: Ternary complex formation of pVHL, elongin B and elongin C visualized in living cells by a fluorescence resonance energy transfer–fluorescence lifetime imaging microscopy technique

The tumor suppressor von Hippel–Lindau (VHL) gene product forms a com-plex with elongin B and elongin C, and acts as a recognition subunit of a ubiquitin E3 ligase. Interactions between components in the complex were investigated in living cells by fluorescence resonance energy transfer (FRET)–fluorescence lifetime imaging microscopy (FLIM). | ỊFEBS Journal Ternary complex formation of pVHL elongin B and elongin C visualized in living cells by a fluorescence resonance energy transfer-fluorescence lifetime imaging microscopy technique Koshi Kinoshita1 Kenji Goryo1 Mamiko Takada2 Yosuke Tomokuni1 Teijiro Aso3 Heiwa Okuda4 Taro Shuin4 Hiroshi Fukumura2 and Kazuhiro Sogawa1 1 Department of Biomolecular Sciences Graduate Schoolof Life Sciences Tohoku University Aoba-ku Sendai Japan 2 Department of Chemistry Graduate Schoolof Science Tohoku University Aoba-ku Sendai Japan 3 Department of FunctionalGenomics Kochi Medical School Kohasu Okoh-cho Nankoku Kochi Japan 4 Department of Urology Kochi Medical School Kohasu Okoh-cho Nankoku Kochi Japan Keywords conformation change FRET-FLIM live cell imaging protein complex ubiquitin ligase Correspondence K. Sogawa Department of Biomolecular Science Graduate School of Life Sciences Tohoku University Aoba-ku Sendai 980-8578 Japan Fax 81 22 795 6594 Tel 81 22 795 6590 E-mail sogawa@ These authors contributed equally to this work Received 26 June 2007 revised 21 August 2007 accepted 29 August 2007 doi The tumor suppressor von Hippel-Lindau VHL gene product forms a complex with elongin B and elongin C and acts as a recognition subunit of a ubiquitin E3 ligase. Interactions between components in the complex were investigated in living cells by fluorescence resonance energy transfer FRET -fluorescence lifetime imaging microscopy FLIM . Elongin B-ceru-lean or cerulean-elongin B was coexpressed with elongin C-citrine or citrine-elongin C in CHO-K1 cells. FRET signals were examined by measuring a change in the fluorescence lifetime of donors and by monitoring a corresponding fluorescence rise of acceptors. Clear FRET signals between elon-gin B and elongin C were observed in all combinations except for the combination of elongin B-cerulean and citrine-elongin C. Although similar experiments to examine interaction between .

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