Photo-induced peptide cleavage in the green-to-red conversion of a fluorescent protein

Hideaki Mizuno, Tapas Kumar Mal, Kit I. Tong, Ryoko Ando, Toshiaki Furuta, Mitsuhiko Ikura, Atsushi Miyawaki

Research output: Contribution to journalArticlepeer-review

257 Scopus citations


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, His62-Tyr63-Gly64, 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 α carbon (Cα) at His62 via a formal β-elimination reaction, which requires the whole, intact protein for its catalysis. The subsequent formation of a double bond between His62-Cα and -Cβ extends the π-conjugation to the imidazole ring of His62, 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.

Original languageEnglish (US)
Pages (from-to)1051-1058
Number of pages8
JournalMolecular cell
Issue number4
StatePublished - Oct 2003

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology


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