Intramolecular dynamics in the environment of the single tryptophan residue in staphylococcal nuclease

Alexander P. Demchenko, Ignacy Gryczynski, Zygmunt Gryczynski, Wieslaw Wiczk, Henryk Malak, Mayer Fishman

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The dipole relaxational dynamics in the environment of a single tryptophan residue Trp-140 in staphylococcal nuclease was studied by time-resolved (multi-frequency phase-modulation) spectroscopy and selective red-edge excitation. The long-wavelength position of the fluorescence spectrum (at 343 nm) and the absence of red-edge excitation effects at 0 and 20°C indicate that this residue is surrounded by very mobile protein groups which relax on the subnanosecond time scale. For these temperatures (0-20°C) the steady-state emission spectra did not show the excitation-wavelength dependent shifts (red-edge effects) for excitation wavelengths from 295 to 308 nm; however, the anisotropy decay rate is slow (tens of nanoseconds). This suggests that the spectral relaxation is due to mobility of the surrounding groups rather than the motion of the tryptophan itself. The motions of the tryptophan surrounding are substantially retarded at reduced temperatures in viscous solvent (60% glycerol). The temperature dependence of the difference in position of fluorescence spectra at excitation wavelengths 295 and 305 nm demonstrate the existence of red-edge effect at sub-zero temperatures, reaching a maximum value at -50°C, where the steady-state emission spectrum is shifted to 332 nm. The excitation and emission wavelength dependence of multi-frequency phase-modulation data at the half-transition point (-40°C) demonstrates the existence of the nanosecond dipolar relaxations. At -40°C the time-dependent spectral shift is close to monoexponential with the relaxation time of 1.4 ns.

Original languageEnglish
Pages (from-to)39-48
Number of pages10
JournalBiophysical Chemistry
Volume48
Issue number1
DOIs
StatePublished - Nov 1993

Fingerprint

Micrococcal Nuclease
Tryptophan
Wavelength
Temperature
Phase modulation
Frequency modulation
Fluorescence
Anisotropy
Glycerol
Spectrum Analysis
Relaxation time
Spectroscopy
Proteins

Keywords

  • Intramolecular dynamics
  • Staphylococcal nuclease
  • Time-resolved fluorescence
  • Tryptophan

Cite this

Demchenko, Alexander P. ; Gryczynski, Ignacy ; Gryczynski, Zygmunt ; Wiczk, Wieslaw ; Malak, Henryk ; Fishman, Mayer. / Intramolecular dynamics in the environment of the single tryptophan residue in staphylococcal nuclease. In: Biophysical Chemistry. 1993 ; Vol. 48, No. 1. pp. 39-48.
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Intramolecular dynamics in the environment of the single tryptophan residue in staphylococcal nuclease. / Demchenko, Alexander P.; Gryczynski, Ignacy; Gryczynski, Zygmunt; Wiczk, Wieslaw; Malak, Henryk; Fishman, Mayer.

In: Biophysical Chemistry, Vol. 48, No. 1, 11.1993, p. 39-48.

Research output: Contribution to journalArticle

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T1 - Intramolecular dynamics in the environment of the single tryptophan residue in staphylococcal nuclease

AU - Demchenko, Alexander P.

AU - Gryczynski, Ignacy

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N2 - The dipole relaxational dynamics in the environment of a single tryptophan residue Trp-140 in staphylococcal nuclease was studied by time-resolved (multi-frequency phase-modulation) spectroscopy and selective red-edge excitation. The long-wavelength position of the fluorescence spectrum (at 343 nm) and the absence of red-edge excitation effects at 0 and 20°C indicate that this residue is surrounded by very mobile protein groups which relax on the subnanosecond time scale. For these temperatures (0-20°C) the steady-state emission spectra did not show the excitation-wavelength dependent shifts (red-edge effects) for excitation wavelengths from 295 to 308 nm; however, the anisotropy decay rate is slow (tens of nanoseconds). This suggests that the spectral relaxation is due to mobility of the surrounding groups rather than the motion of the tryptophan itself. The motions of the tryptophan surrounding are substantially retarded at reduced temperatures in viscous solvent (60% glycerol). The temperature dependence of the difference in position of fluorescence spectra at excitation wavelengths 295 and 305 nm demonstrate the existence of red-edge effect at sub-zero temperatures, reaching a maximum value at -50°C, where the steady-state emission spectrum is shifted to 332 nm. The excitation and emission wavelength dependence of multi-frequency phase-modulation data at the half-transition point (-40°C) demonstrates the existence of the nanosecond dipolar relaxations. At -40°C the time-dependent spectral shift is close to monoexponential with the relaxation time of 1.4 ns.

AB - The dipole relaxational dynamics in the environment of a single tryptophan residue Trp-140 in staphylococcal nuclease was studied by time-resolved (multi-frequency phase-modulation) spectroscopy and selective red-edge excitation. The long-wavelength position of the fluorescence spectrum (at 343 nm) and the absence of red-edge excitation effects at 0 and 20°C indicate that this residue is surrounded by very mobile protein groups which relax on the subnanosecond time scale. For these temperatures (0-20°C) the steady-state emission spectra did not show the excitation-wavelength dependent shifts (red-edge effects) for excitation wavelengths from 295 to 308 nm; however, the anisotropy decay rate is slow (tens of nanoseconds). This suggests that the spectral relaxation is due to mobility of the surrounding groups rather than the motion of the tryptophan itself. The motions of the tryptophan surrounding are substantially retarded at reduced temperatures in viscous solvent (60% glycerol). The temperature dependence of the difference in position of fluorescence spectra at excitation wavelengths 295 and 305 nm demonstrate the existence of red-edge effect at sub-zero temperatures, reaching a maximum value at -50°C, where the steady-state emission spectrum is shifted to 332 nm. The excitation and emission wavelength dependence of multi-frequency phase-modulation data at the half-transition point (-40°C) demonstrates the existence of the nanosecond dipolar relaxations. At -40°C the time-dependent spectral shift is close to monoexponential with the relaxation time of 1.4 ns.

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