GrpE accelerates peptide binding and release from the high affinity state of DnaK.

The Escherichia coli nucleotide exchange factor GrpE accelerates the rate of ADP dissociation from high affinity ADP-DnaK, thus enabling ATP binding and transition to the low affinity state. We show here that GrpE, in the absence of ATP, accelerates the rates of the forward and reverse reaction ADP-DnaK-P right harpoon over left harpoon ADP-DnaK + P, where P denotes peptide substrate. Specifically, the binding of GrpE to an ADP-DnaK-P (or DnaK-P) complex increases koff and kon by approximately 200-fold and approximately 60-fold, respectively. The results are consistent with a GrpE- induced conformational change in the C-terminal polypeptide binding domain of an ADP-DnaK molecule, which results in a unique low affinity intermediate from which peptide can dissociate. A simulation of peptide dissociation from DnaK as a function of the [ATP] / [ADP] ratio shows that GrpE induced peptide dissociation from ADP-DnaK is important at elevated cellular concentrations of ADP, which typically occur upon stress.