We investigated the folding of the 70 kDa human cytosolic inducible protein (Hsp70) in vitro using high hydrostatic pressure as a denaturing agent. We followed the structural changes in Hsp70 induced by high hydrostatic pressure using tryptophan fluorescence, molecular dynamics, circular dichroism, HPLC gel filtration, dynamic light scattering, ATPase activity and chaperone activity. Although monomeric, Hsp70 is very sensitive to hydrostatic pressure; after pressure removal, the protein did not return to its native sate but instead formed oligomeric species that lost chaperone activity but retained ATPase activity.