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Changing times: Fluorescence-lifetime analysis of amyloidogenic SF-IAPP fusion protein.
Antimonova, Olga I; Lebedev, Dmitry V; Zabrodskaya, Yana A; Grudinina, Natalia A; Timkovsky, Andrey L; Ramsay, Edward; Shavlovsky, Michael M; Egorov, Vladimir V.
Afiliación
  • Antimonova OI; Department of Molecular Genetics, Federal State Budgetary Scientific Institution "Institute of Experimental Medicine", 197376 Akademika Pavlova St. 12, St. Petersburg, Russia.
  • Lebedev DV; Department of Molecular and Radiation Biophysics, Petersburg Nuclear Physics Institute named by B. P. Konstantinov of National Research Center "Kurchatov Institute", 188300 mkr. Orlova roshcha 1, Gatchina, Russia.
  • Zabrodskaya YA; Department of Molecular and Radiation Biophysics, Petersburg Nuclear Physics Institute named by B. P. Konstantinov of National Research Center "Kurchatov Institute", 188300 mkr. Orlova roshcha 1, Gatchina, Russia; Department of Molecular Biology of Viruses, Smorodintsev Research Institute of Influen
  • Grudinina NA; Department of Molecular Genetics, Federal State Budgetary Scientific Institution "Institute of Experimental Medicine", 197376 Akademika Pavlova St. 12, St. Petersburg, Russia.
  • Timkovsky AL; Department of Molecular and Radiation Biophysics, Petersburg Nuclear Physics Institute named by B. P. Konstantinov of National Research Center "Kurchatov Institute", 188300 mkr. Orlova roshcha 1, Gatchina, Russia; Peter the Great St. Petersburg Polytechnic University, 195251 Polytechnicheskaya St. 2
  • Ramsay E; Department of Molecular Biology of Viruses, Smorodintsev Research Institute of Influenza, Ministry of Healthcare of the Russian Federation, 197376 Prof. Popov St. 15/17, St. Petersburg, Russia.
  • Shavlovsky MM; Department of Molecular Genetics, Federal State Budgetary Scientific Institution "Institute of Experimental Medicine", 197376 Akademika Pavlova St. 12, St. Petersburg, Russia; North-Western State Medical University named after I.I. Mechnikov, 191015 Kirochnaya St. 41, St. Petersburg, Russia.
  • Egorov VV; Department of Molecular Genetics, Federal State Budgetary Scientific Institution "Institute of Experimental Medicine", 197376 Akademika Pavlova St. 12, St. Petersburg, Russia; Department of Molecular and Radiation Biophysics, Petersburg Nuclear Physics Institute named by B. P. Konstantinov of Nation
J Struct Biol ; 205(1): 78-83, 2019 01 01.
Article en En | MEDLINE | ID: mdl-30458241
In a number of conformational diseases, intracellular accumulation of proteins bearing non-native conformations occurs. The search for compounds that are capable of hindering the formation and accumulation of toxic protein aggregates and fibrils is an urgent task. Present fluorescent methods of fibrils' detection prevent simple real-time observations. We suppose to use green fluorescent protein fused with target protein and fluorescence lifetime measurement technique for this purpose. The recombinant proteins analyzed were produced in E. coli. Mass spectrometry was used for the primary structure of the recombinant proteins and post-translational modifications identification. The fluorescence lifetime of the superfolder green fluorescent protein (SF) and the SF protein fused with islet amyloid polypeptide (SF-IAPP) were studied in polyacrylamide gel using Fluorescent-Lifetime Imaging Microscopy (FLIM). It was shown that the SF average fluorescence lifetime in gel slightly differs from that of the SF-IAPP monomer under these conditions. SF-IAPP does not lose the ability to form amyloid-like fibrils. Under the same conditions (in polyacrylamide gel), SF and SF-IAPP monomers have similar fluorescence time characteristics and the average fluorescence lifetime of SF-IAPP in fibrils significantly decreases. We propose the application of FLIM to the measurement of average fluorescence lifetimes of fusion proteins (amyloidogenic protein-SF) in the context of studies using cellular models of conformational diseases.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Proteínas Recombinantes / Proteínas Fluorescentes Verdes / Polipéptido Amiloide de los Islotes Pancreáticos / Imagen Óptica Tipo de estudio: Prognostic_studies Límite: Animals / Humans Idioma: En Revista: J Struct Biol Asunto de la revista: BIOLOGIA MOLECULAR Año: 2019 Tipo del documento: Article País de afiliación: Rusia Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Proteínas Recombinantes / Proteínas Fluorescentes Verdes / Polipéptido Amiloide de los Islotes Pancreáticos / Imagen Óptica Tipo de estudio: Prognostic_studies Límite: Animals / Humans Idioma: En Revista: J Struct Biol Asunto de la revista: BIOLOGIA MOLECULAR Año: 2019 Tipo del documento: Article País de afiliación: Rusia Pais de publicación: Estados Unidos