INHIBITION OF DNAK AUTOPHOSPHORYLATION BY HEAT-SHOCK PROTEINS AND POLYPEPTIDE SUBSTRATES

DnaK, the Hsp70 of Escherichia coil, autophosphorylates in vitro. Of t he two heat shock proteins that interact with DnaK, GrpE inhibits DnaK phosphorylation, whereas DnaJ has no effect on the reaction. Three sy nthetic peptides are shown to inhibit DnaK phosphorylation. The potenc y of a given peptide correlates with its affinity for the DnaK protein . A truncated DnaK that lacks the carboxyl-terminal peptide-binding do main autophosphorylates; this reaction is resistant to the inhibitory peptides. Phosphorylation of the truncated DnaK is still inhibited by GrpE, indicating that the GrpE-binding site resides in the DnaK amino- terminal domain. Thus, DnaK phosphorylation is regulated in vitro, and possibly in vivo, by physiologically relevant substrates and cofactor s.