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Fluorescence Quenching Studies on Enzyme I of the Bacterial Phosphotransferase System

  • Paolo Neyroz
  • Bonnie Bassler
  • Ludwig Brand
  • Saul Roseman
Conference paper

Abstract

Enzyme I is the first protein involved in the series of phosphoryl transfer steps catalyzed by the glycose: pyruvate phosphotransferase system (PTS) (Meadow et al., 1984). The PTS is responsible for the phosphoryla-tion and concomitant transport of its sugar substrates across the bacterial membrane, and in this process phosphoenolpyruvate (PEP) serves as the phosphoryl donor. It has been previously observed (Kukuruzinska et al., 1982) that the subunits of Enzyme I associate in a temperature-dependent manner to form dimers. The enzyme contains two tryptophans per subunit and their intrinsic emission have been characterized at 2°C for the monomer and 23°C for the dimer. The decay of the fluorescence intensity has been resolved into a double exponential decay for both the conformations of the protein (Neyroz et al., 1984). At 2°C a short lifetime of 3 ns and a long one of 7.5 ns were found that account for the 17% and 83% of the total emitted light, respectively. At 23°C the recovered decay constants were 3.7 ns and 7.5 ns with an increase of the fractional contribution of the short lifetime to 43% of the total intensity. Two different decay associated spectra (DAS) (Knutson et al., 1982) were obtained for each decay component. A spectrum with its maximum at 330 nm was found for the short lifetime and a second spectrum centered at 345 nm was resolved for the longer lifetime. Changes in the proportions of the two spectra obtained at 2°C and 23°C well reproduced the difference of amplitudes reported above when the protein was excited at 295 nm and the fluorescence observed at 340 nm.

Keywords

Fluorescence Lifetime Tryptophan Residue Short Lifetime Fluorescence Decay Decay Component 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Plenum Press, New York 1989

Authors and Affiliations

  • Paolo Neyroz
    • 1
  • Bonnie Bassler
    • 1
  • Ludwig Brand
    • 1
  • Saul Roseman
    • 1
  1. 1.Department of BiologyThe Johns Hopkins UniversityBaltimoreUSA

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