Comparative studies on the structure and stability of fluorescent proteins EGFP, zFP506, mRFP1, "dimer2", and DsRed1

Olesia V. Stepanenko, Vladislav V. Verkhusha, Vasili I. Kazakov, Michail M. Shavlovsky, Irina M. Kuznetsova, Vladimir N. Uversky, Konstantin K. Turoverov

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Abstract

To obtain more information about the structural properties and conformational stabilities of GFP-like fluorescent proteins, we have undertaken a systematic analysis of series of green and red fluorescent proteins with different association states. The list of studied proteins includes EGFP (green monomer), zFP506 (green tetramer), mRFP1 (red monomer), "dimer2" (red dimer), and DsRed1 (red tetramer). Fluorescent and absorbance parameters, near-UV and visible CD spectra, the accessibility of the chromophores and tryptophans to acrylamide quenching, and the resistance of these proteins to the guanidine hydrochloride unfolding and kinetics of the approaching of the unfolding equilibrium have been compared. Tetrameric zFP506 was shown to be dramatically more stable than the EGFP monomer, assuming that association might contribute to the protein conformational stability. This assumption is most likely valid even though the sequences OF GFP and zPF506 are only ∼25% identical. Interestingly, red FPs possessed comparable conformational stabilities, where monomeric mRFP1 was the most stable species under the equilibrium conditions, whereas the tetrameric DsRed1 possessed the slowest unfolding kinetics. Furthermore, EGFP is shown to be considerably less stable than mRFP1, whereas tetrameric zFP506 is the most stable species analyzed in this study. This means that the quaternary structure, being an important stabilizing factor, does not represent the only circumstance dictating the dramatic variations between fluorescent proteins in their conformational stabilities.

Original languageEnglish (US)
Pages (from-to)14913-14923
Number of pages11
JournalBiochemistry
Volume43
Issue number47
DOIs
Publication statusPublished - Nov 30 2004
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry

Cite this

Stepanenko, O. V., Verkhusha, V. V., Kazakov, V. I., Shavlovsky, M. M., Kuznetsova, I. M., Uversky, V. N., & Turoverov, K. K. (2004). Comparative studies on the structure and stability of fluorescent proteins EGFP, zFP506, mRFP1, "dimer2", and DsRed1. Biochemistry, 43(47), 14913-14923. https://doi.org/10.1021/bi048725t