A colourless green fluorescent protein homologue from the non-fluorescent hydromedusa Aequorea coerulescens and its fluorescent mutants

Nadya G. Gurskaya, Arkady F. Fradkov, Natalia I. Pounkova, Dmitry B. Staroverov, Maria E. Bulina, Yurii G. Yanushevich, Yulii A. Labas, Sergey Lukyanov, Konstantin A. Lukyanov

Research output: Contribution to journalArticlepeer-review

72 Citations (Scopus)

Abstract

We have cloned an unusual colourless green fluorescent protein (GFP)-like protein from Aequorea coerulescens (acGFPL). The A. coerulescens specimens displayed blue (not green) luminescence, and no fluorescence was detected in these medusae. Escherichia coli expressing wild-type acGFPL showed neither fluorescence nor visible coloration. Random mutagenesis generated green fluorescent mutants of acGFPL, with the strongest emitters found to contain an Glu222 → Gly (E222G) substitution, which removed the evolutionarily invariant Glu222. Reintroduction of Glu222 into the most fluorescent random mutant, named aceGFP, converted it into a colourless protein. This colourless aceGFP-G222E protein demonstrated a novel type of UV-induced photoconversion, from an immature non-fluorescent form into a green fluorescent form. Fluorescent aceGFP may be a useful biological tool, as it was able to be expressed in a number of mammalian cell lines. Furthermore, expression of a fusion protein of 'humanized' aceGFP and β-actin produced a fluorescent pattern consistent with actin distribution in mammalian cells.

Original languageEnglish
Pages (from-to)403-408
Number of pages6
JournalBiochemical Journal
Volume373
Issue number2
DOIs
Publication statusPublished - 15 Jul 2003
Externally publishedYes

Keywords

  • Bioluminescence
  • Green fluorescent protein (GFP)
  • Photoconversion
  • Protein labelling

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