Heat perturbation of bovine eye lens α-crystallin probed by covalently attached ratiometric fluorescent dye 4′-diethylamino-3- hydroxyflavone

S. V. Avilov, Cs Bode, F. Tölgyesi, A. S. Klymchenko, J. Fidy, A. P. Demchenko

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Bovine eye lens α-crystallin was covalently labeled with 6-bromomethyl-4′-diethylamino-3-hydroxyflavone and studied under native-like conditions and at the elevated temperature (60°C) that is known to facilitate α-crystallin chaperone-like activity. This novel SH-reactive two-band ratiometric fluorescent probe is characterized by two highly emissive N*- and T*-bands; the latter appears due to excited state intramolecular proton transfer reaction. The positions of these bands and the ratio of their intensities for the α-crystallin-dye conjugate are the sensitive indicators of polarity of the dye environment and its participation in intermolecular hydrogen bonding. Although we found that the dye labels both the SH and the NH2 groups in α-crystallin, a recently developed procedure allowed us to distinguish between the heat-induced spectral changes of the dye molecules attached to SH and NH2 groups. We observed that at elevated temperature the environment of the SH-attached dye becomes more polar and flexible. The number of H-bond acceptor groups in the vicinity of the dye decreases. Since α-crystallin contains a single Cys residue within the C-terminal domain of its αA subunit (the αB subunit contains none), we can attribute the observed effects to temperature-induced changes in the C-terminal domain of this protein.

Original languageEnglish
Pages (from-to)340-348
Number of pages9
JournalBiopolymers - Peptide Science Section
Volume78
Issue number6
DOIs
Publication statusPublished - Aug 15 2005

Fingerprint

Crystalline Lens
Crystallins
Fluorescent Dyes
Lenses
Coloring Agents
Dyes
Hot Temperature
Temperature
Proton transfer
Hydrogen Bonding
Excited states
4'-diethylamino-3-hydroxyflavone
Protons
Labels
Hydrogen bonds
Proteins
Molecules

Keywords

  • 6-bromomethyl-4′-diethylamino-3-hydroxyflavone
  • Eye lens α-crystallin
  • Ratiometric fluorescence probe
  • Small heat-shock protein
  • Temperature perturbation

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biophysics

Cite this

Heat perturbation of bovine eye lens α-crystallin probed by covalently attached ratiometric fluorescent dye 4′-diethylamino-3- hydroxyflavone. / Avilov, S. V.; Bode, Cs; Tölgyesi, F.; Klymchenko, A. S.; Fidy, J.; Demchenko, A. P.

In: Biopolymers - Peptide Science Section, Vol. 78, No. 6, 15.08.2005, p. 340-348.

Research output: Contribution to journalArticle

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