BAG-1, a negative regulator of Hsp70 chaperone activity, uncouples nucleotide hydrolysis from substrate release.
Molecular chaperones influence the process of protein folding and, under conditions of stress, recognize non-native proteins to ensure that misfolded proteins neither appear nor accumulate. BAG-1, identified as an Hsp70 associated protein, was shown to have the unique properties of a negative regulator of Hsp70. Here, we demonstrate that BAG-1 inhibits the in vitro protein refolding activity of Hsp70 by forming stable ternary complexes with non-native substrates that do not release even in the presence of nucleotide and the co-chaperone, Hdj-1. However, the substrate in the BAG-1-containing ternary complex does not aggregate and remains in a soluble intermediate folded state, indistinguishable from the refolding-competent substrate-Hsp70 complex. BAG-1 neither inhibits the Hsp70 ATPase, nor has the properties of a nucleotide exchange factor; instead, it stimulates ATPase activity, similar to that observed for Hdj-1, but with opposite consequences. In the presence of BAG-1, the conformation of Hsp70 is altered such that the substrate binding domain becomes less accessible to protease digestion, even in the presence of nucleotide and Hdj-1. These results suggest a mechanistic basis for BAG-1 as a negative regulator of the Hsp70-Hdj-1 chaperone cycle.
Bimston
D
D
Department of Biochemistry, Molecular Biology and Cell Biology, Rice Institute for Biomedical Research, Northwestern University, Evanston, IL 60208, USA.
Song
J
J
Winchester
D
D
Takayama
S
S
Reed
J C
JC
Morimoto
R I
RI
D000251:Adenosine Triphosphatases / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D004268:DNA-Binding Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D011487:Protein Conformation; D013379:Substrate Specificity; D014157:Transcription Factors
C091732:BCL2-associated athanogene 1 protein; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D014157:Transcription Factors; D000251:Adenosine Triphosphatases