J Proteins Catalytically Activate Hsp70 Molecules to Trap a Wide Range of Peptide Sequences

Howard Hughes Medical Institute, Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Molecular Cell (Impact Factor: 14.46). 12/1998; 2(5):593-603. DOI: 10.1016/S1097-2765(00)80158-6
Source: PubMed

ABSTRACT Proteins of the Hsp70 family of ATPases, such as BiP, function together with J proteins to bind polypeptides in numerous cellular processes. Using a solid phase binding assay, we demonstrate that a conserved segment of the J proteins, the J domain, catalytically activates BiP molecules to bind peptides in its immediate vicinity. The J domain interacts with the ATP form of BiP and stimulates hydrolysis resulting in the rapid trapping of peptides, which are then only slowly released upon nucleotide exchange. Activation by the J domain allows BiP to trap peptides or proteins that it would not bind on its own. These results explain why BiP and probably all other Hsp70s can interact with a wide range of substrates and suggest that the J partner primarily determines the substrate specificity of Hsp70s.

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    • "We have assumed that a single BiP is activated per Sec63 molecule located at the ER membrane. In vitro studies have indicated that the BiP-Sec63 interaction occurs transiently, and suggest that one Sec63 molecule could potentially activate at least ten BiP molecules [38]. On average, E. coli experiments have determined that incoming proteins present a new Hsp70 binding site every 25 to 35 amino acids [46] which is consistent with S. cerevisiae literature that estimates a minimum of six to seven molecules of BiP bound to the endogeneous protein, ppαF [47]. "
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    • "Second, GRP78 knockdown did not affect N-linked glycosylation, supporting that IP 3 R1 is normally folded by the ER lectin chaperone calnexin (Joseph et al., 1999; Ellgaard and Helenius, 2003) and that IP 3 R1 does not undergo disassembly-dependent degradation (Khan and Joseph, 2003). Third, the specific binding behavior of GRP78 to IP 3 R1 is incompatible with typical binding behaviors of GRP78 to unfolded substrates regarding the dissociation kinetics and the preferences of ATP and ADP (Misselwitz et al., 1998; Hendershot, 2004; Nawa et al., 2007). Therefore, our findings strongly support the argument that GRP78 functions as an assembler of IP 3 R1. "
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    • "For example, while Ssa1/2 have general protein folding functions in the cytosol, Sis1 assists Ssa1/2 on ribosomes in translation [59], Ydj1 recruits Ssa1/2 to protein folding on ER membranes [61] and Djp1 stimulates Ssa1/2 to assist in peroxisome biogenesis [58]. Interaction of Hsp40 with various substrates as well as its ability to stimulate the Hsp70 reaction cycle allows Hsp70 to trap a wider range of substrates than it would be capable of binding on its own [62]. "
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