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Modulation of Ultrafast Conformational Dynamics in Allosteric Interaction of Gal Repressor Protein with Different Operator DNA Sequences
Author(s) -
Choudhury Susobhan,
Naiya Gitashri,
Singh Priya,
Lemmens Peter,
Roy Siddhartha,
Pal Samir Kumar
Publication year - 2016
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201500657
Subject(s) - allosteric regulation , förster resonance energy transfer , chemistry , protein dynamics , biophysics , lac repressor , allosteric enzyme , cooperative binding , repressor , cooperativity , protein structure , fluorescence , binding site , biochemistry , biology , physics , transcription factor , quantum mechanics , gene , enzyme
Although all forms of dynamical behaviour of a protein under allosteric interaction with effectors are predicted, little evidence of ultrafast dynamics in the interaction has been reported. Here, we demonstrate the efficacy of a combined approach involving picosecond‐resolved FRET and polarisation‐gated fluorescence for the exploration of ultrafast dynamics in the allosteric interaction of the Gal repressor (GalR) protein dimer with DNA operator sequences O E and O I . FRET from the single tryptophan residue to a covalently attached probe IAEDANS at a cysteine residue in the C‐terminal domain of GalR shows structural perturbation and conformational dynamics during allosteric interaction. Polarisation‐gated fluorescence spectroscopy of IAEDANS and another probe (FITC) covalently attached to the operator directly revealed the essential dynamics for cooperativity in the protein–protein interaction. The ultrafast resonance energy transfer from IAEDANS in the protein to FITC also revealed different dynamic flexibility in the allosteric interaction. An attempt was made to correlate the dynamic changes in the protein dimers with O E and O I with the consequent protein–protein interaction (tetramerisation) to form a DNA loop encompassing the promoter segment.