Photo-induced peptide cleavage in the green-to-red conversion of a fluorescent protein |
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Authors: | Mizuno Hideaki Mal Tapas Kumar Tong Kit I Ando Ryoko Furuta Toshiaki Ikura Mitsuhiko Miyawaki Atsushi |
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Institution: | Laboratory for Cell Function Dynamics, Advanced Technology Development Group, Brain Science Institute, The Institute of Physical and Chemical Science (RIKEN), 2-1 Hirosawa, Wako-city, Saitama 351-0198, Japan. |
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Abstract: | Green fluorescent protein from the jellyfish (Aequorea GFP) and GFP-like proteins from coral species encode light-absorbing chromophores within their protein sequences. A coral fluorescent protein, Kaede, contains a tripeptide, His(62)-Tyr(63)-Gly(64), which acts as a green chromophore that is photoconverted to red. Here, we present the structural basis for the green-to-red photoconversion. As in Aequorea GFP, a chromophore, 4-(p-hydroxybenzylidene)-5-imidazolinone, derived from the tripeptide mediates green fluorescence in Kaede. UV irradiation causes an unconventional cleavage within Kaede protein between the amide nitrogen and the alpha carbon (Calpha) at His(62) via a formal beta-elimination reaction, which requires the whole, intact protein for its catalysis. The subsequent formation of a double bond between His(62)-Calpha and -Cbeta extends the pi-conjugation to the imidazole ring of His(62), creating a new red-emitting chromophore, 2-(1E)-2-(5-imidazolyl)ethenyl]-4-(p-hydroxybenzylidene)-5-imidazolinone. The present study not only reveals diversity in the chemical structure of fluorescent proteins but also adds a new dimension to posttranslational modification mechanisms. |
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