Conformational distributions of melittin in water / methanol mixtures from frequency-domain measurements of nonradiative energy transfer

Joseph R. Lakowicz, Ignacy Gryczynski, Wieslaw Wiczk, Gabor Laczko, Franklyn C. Prendergast, Michael L. Johnson

Research output: Contribution to journalArticle

63 Citations (Scopus)

Abstract

We used fluorescence energy transfer to examine the effects of solvent composition on the distribution of distances between the single tryptophan residue of melittin (residue 19) to the N-terminal α-amino group, which was labeled with a dansyl residue. The tryptophan intensity decays, with and without the dansyl acceptor, were measured by the frequency-domain method. The data were analyzed by a least-squares algorithm which accounts for correlation between the parameters. A wide distribution of tryptophan to dansyl distances was found for the random-coil state, with a Gaussian half-width of 25 Å. Increasing concentrations of methanol, which were shown to induce an α-helical conformation, resulted in a progressive decrease in the width of the distribution, reaching a limiting half-width of 3 Å at 80% (v/v) methanol. The distance from the indole moiety of Trp-19 to the dansyl group in 80% (v/v) methanol/water was found to be 25 Å, as assessed from the center of the distance distribution. A distance of 24-25 Å was recovered from the X-ray crystal structure of the tetramer, which is largely α-helical. At low ionic strength ( < 0.01) the CD spectra revealed a small fraction or amount of α-helix for molittin in water, which implies a small fraction of residual structure. This residual structure is apparently lost in guanidine hydrochloride as demonstrated by a further broadening in the distribution of distances. These results demonstrate the usefulness of frequency-domain measurements of resonance transfer for resolution of conformational distributions of proteins.

Original languageEnglish (US)
Pages (from-to)99-115
Number of pages17
JournalBiophysical Chemistry
Volume36
Issue number2
DOIs
StatePublished - 1990

Fingerprint

Melitten
Energy Transfer
Tryptophan
Energy transfer
Methanol
methyl alcohol
energy transfer
Water
tryptophan
water
Guanidine
Ionic strength
Least-Squares Analysis
Osmolar Concentration
Conformations
Crystal structure
Fluorescence
X-Rays
guanidines
X rays

Keywords

  • Distance distribution
  • Energy transfer
  • Fluorescence spectroscopy
  • Frequency-domain fluorescence
  • Melittin
  • Time-resolved fluorescence

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Physical and Theoretical Chemistry

Cite this

Conformational distributions of melittin in water / methanol mixtures from frequency-domain measurements of nonradiative energy transfer. / Lakowicz, Joseph R.; Gryczynski, Ignacy; Wiczk, Wieslaw; Laczko, Gabor; Prendergast, Franklyn C.; Johnson, Michael L.

In: Biophysical Chemistry, Vol. 36, No. 2, 1990, p. 99-115.

Research output: Contribution to journalArticle

Lakowicz, Joseph R. ; Gryczynski, Ignacy ; Wiczk, Wieslaw ; Laczko, Gabor ; Prendergast, Franklyn C. ; Johnson, Michael L. / Conformational distributions of melittin in water / methanol mixtures from frequency-domain measurements of nonradiative energy transfer. In: Biophysical Chemistry. 1990 ; Vol. 36, No. 2. pp. 99-115.
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abstract = "We used fluorescence energy transfer to examine the effects of solvent composition on the distribution of distances between the single tryptophan residue of melittin (residue 19) to the N-terminal α-amino group, which was labeled with a dansyl residue. The tryptophan intensity decays, with and without the dansyl acceptor, were measured by the frequency-domain method. The data were analyzed by a least-squares algorithm which accounts for correlation between the parameters. A wide distribution of tryptophan to dansyl distances was found for the random-coil state, with a Gaussian half-width of 25 {\AA}. Increasing concentrations of methanol, which were shown to induce an α-helical conformation, resulted in a progressive decrease in the width of the distribution, reaching a limiting half-width of 3 {\AA} at 80{\%} (v/v) methanol. The distance from the indole moiety of Trp-19 to the dansyl group in 80{\%} (v/v) methanol/water was found to be 25 {\AA}, as assessed from the center of the distance distribution. A distance of 24-25 {\AA} was recovered from the X-ray crystal structure of the tetramer, which is largely α-helical. At low ionic strength ( < 0.01) the CD spectra revealed a small fraction or amount of α-helix for molittin in water, which implies a small fraction of residual structure. This residual structure is apparently lost in guanidine hydrochloride as demonstrated by a further broadening in the distribution of distances. These results demonstrate the usefulness of frequency-domain measurements of resonance transfer for resolution of conformational distributions of proteins.",
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N2 - We used fluorescence energy transfer to examine the effects of solvent composition on the distribution of distances between the single tryptophan residue of melittin (residue 19) to the N-terminal α-amino group, which was labeled with a dansyl residue. The tryptophan intensity decays, with and without the dansyl acceptor, were measured by the frequency-domain method. The data were analyzed by a least-squares algorithm which accounts for correlation between the parameters. A wide distribution of tryptophan to dansyl distances was found for the random-coil state, with a Gaussian half-width of 25 Å. Increasing concentrations of methanol, which were shown to induce an α-helical conformation, resulted in a progressive decrease in the width of the distribution, reaching a limiting half-width of 3 Å at 80% (v/v) methanol. The distance from the indole moiety of Trp-19 to the dansyl group in 80% (v/v) methanol/water was found to be 25 Å, as assessed from the center of the distance distribution. A distance of 24-25 Å was recovered from the X-ray crystal structure of the tetramer, which is largely α-helical. At low ionic strength ( < 0.01) the CD spectra revealed a small fraction or amount of α-helix for molittin in water, which implies a small fraction of residual structure. This residual structure is apparently lost in guanidine hydrochloride as demonstrated by a further broadening in the distribution of distances. These results demonstrate the usefulness of frequency-domain measurements of resonance transfer for resolution of conformational distributions of proteins.

AB - We used fluorescence energy transfer to examine the effects of solvent composition on the distribution of distances between the single tryptophan residue of melittin (residue 19) to the N-terminal α-amino group, which was labeled with a dansyl residue. The tryptophan intensity decays, with and without the dansyl acceptor, were measured by the frequency-domain method. The data were analyzed by a least-squares algorithm which accounts for correlation between the parameters. A wide distribution of tryptophan to dansyl distances was found for the random-coil state, with a Gaussian half-width of 25 Å. Increasing concentrations of methanol, which were shown to induce an α-helical conformation, resulted in a progressive decrease in the width of the distribution, reaching a limiting half-width of 3 Å at 80% (v/v) methanol. The distance from the indole moiety of Trp-19 to the dansyl group in 80% (v/v) methanol/water was found to be 25 Å, as assessed from the center of the distance distribution. A distance of 24-25 Å was recovered from the X-ray crystal structure of the tetramer, which is largely α-helical. At low ionic strength ( < 0.01) the CD spectra revealed a small fraction or amount of α-helix for molittin in water, which implies a small fraction of residual structure. This residual structure is apparently lost in guanidine hydrochloride as demonstrated by a further broadening in the distribution of distances. These results demonstrate the usefulness of frequency-domain measurements of resonance transfer for resolution of conformational distributions of proteins.

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