Interdomain interaction through helices A and B of DnaK peptide binding domain.

In order to better define the structural elements involved in allosteric signalling, wild-type DnaK and three deletion mutants of the peptide binding domain have been characterized by biophysical (steady-state and time-resolved fluorescence) and biochemical methods. In the presence of ATP the chemical environment of the single tryptophan residue of DnaK, located in the ATPase domain, becomes less polar, as seen by a blue shift of the emission maximum and a shortening of the fluorescence lifetime, and its accessibility to polar quenchers is drastically reduced. These nucleotide-dependent modifications are also observed for the deletion mutant DnaK1-537, but not for DnaK1-507 or DnaK1-385, and thus rely on the presence of residues 507-537 (helices A and the N-terminal half of B) of the peptide binding domain. These data indicate that alphaA and half alphaB contribute to the allosteric communication of DnaK. In the presence of ATP, they promote a conformational change that displaces a residue(s) of the peptide binding domain towards a region of the ATPase domain where the tryptophan residue (W102) is located. A putative role for these helical segments as regulators of the position of the lid is discussed.