The Hsp 70 and Hsp 60 Review Chaperone Machines

In Hsp70-assisted folding reactions, substrates unAn essential cellular machinery that has been identified dergo repeated cycles of binding/release (Szabo et al., and studied only relatively recently is a collective of 1994; Buchberger et al., 1996), frequently at a stoichiomspecialized proteins, molecular chaperones, that bind etry of a single Hsp70 monomer per substrate molecule. nonnative states of other proteins and assist them to Hsp70 binding does not appear to induce global conforreach a functional conformation, in most cases through mational changes in the substrate but, rather, appears the expenditure of ATP. Originally identified by their into act locally. Substrates released from the chaperone creased abundance following heat shock, chaperone undergo kinetic partitioning between folding to native proteins in general recognize exposed hydrophobic surstate, aggregation, rebinding to Hsp70, and binding to faces of nonnative species, surfaces that will ultimately other chaperones or proteases as part of a multidirecbe buried in the native state, and form noncovalent intertional folding network. Hsp70 proteins all consist of the actions with them, stabilizing them against irreversible same working parts: a highly conserved NH2-terminal multimeric aggregation. Release of polypeptide then folATPase domain of 44 kDa and a COOH-terminal region lows, in many cases driven by an ATP-directed conforof 25 kDa, divided into a conserved substrate binding mational change of the chaperone, permitting subsedomain of 15 kDa and a less-conserved immediate quent steps of polypeptide folding or biogenesis to COOH-terminal domain of 10 kDa (Figure 1). occur. When such steps fail to proceed productively, Structure of an Hsp70 Peptide Binding Domain, recognition and rebinding by the same or another chapand the Consensus Motif Recognized erone can occur, allowing another opportunity for a proin Substrate Proteins ductive conformation to be reached. The molecular basis for Hsp70 binding to many nonnaDifferent classes of molecular chaperones appear to tive proteins has been elucidated for the bacterial cybe directed to binding specific nonnative states, the toplasmic homolog, DnaK, through biochemical and nature of which are beginning to be understood. For crystallographic studies. Studies with peptides have inexample, the two best-studied families, examined in dedicated that DnaK binds with greatest affinity to short tail below, theubiquitous Hsp70 and Hsp60 (chaperonin) hydrophobic segments in extended conformation (Schmid chaperones, recognize hydrophobic surface in the conet al., 1994; Zhu et al., 1996). To qualify as a substrate, text of, respectively, extended and collapsed (globular) it thus seems a minimal requirement that a protein exconformations, which are bound correspondingly either pose a single recognizable segment, either through local by local enclosure of the chain or by global enclosure unfolding or as an intrinsically unfolded structural eleof the polypeptide in a central cavity. Because there is ment, such as a loop. The crystal structure of the COOHnot yet the detailed level of structural and mechanistic terminal substrate binding domain of DnaK complexed understanding for other recognized families of chaperwith a heptapeptide substrate, NRLLLTG (Zhu et al., ones, they are not considered here, but important obser1996), reveals a b sandwich composed of two sheets vations concerning binding, nucleotide use, and cellular of four strands each, followed in the primary structure actions are summarized in Table 1. by two a helices, A and B, which span back over the