Bag-1M accelerates nucleotide release for human Hsc70 and Hsp70 and can act concentration-dependent as positive and negative cofactor.
The cytosol of mammalian cells contains several Hsp70 chaperones and an arsenal of cochaperones, including the anti-apoptotic Bag-1M protein, which regulate the activities of Hsp70s by controlling their ATPase cycles. To elucidate the regulatory function of Bag-1M, we determined its influence on nucleotide exchange, substrate release, ATPase rate, and chaperone activity of the housekeeping Hsc70 and stress-inducible Hsp70 homologs of humans. Bag-1M and a C-terminal fragment of it are potent nucleotide exchange factors as they stimulated the ADP dissociation rate of Hsc70 and Hsp70 up to 900-fold. The N-terminal domain of Bag-1M decreased the affinity of Bag-1M for Hsc70/Hsp70 by 4-fold, indicating a modulating role of the N terminus in Bag-1M action as nucleotide exchange factor. Bag-1M inhibited Hsc70/Hsp70-dependent refolding of luciferase in the absence of P(i). Surprisingly, under physiological conditions, i.e. low Bag-1M concentrations and presence of P(i), Bag-1M activates the chaperone action of Hsc70/Hsp70 in luciferase refolding. Bag-1M accelerated ATP-triggered substrate release by Hsc70/Hsp70. We propose that Bag-1M acts as substrate discharging factor for Hsc70 and Hsp70.
Gassler
C S
CS
Institut für Biochemie und Molekularbiologie, Universität Freiburg, 79104 Freiburg, Germany.
Wiederkehr
T
T
Brehmer
D
D
Bukau
B
B
Mayer
M P
MP
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008156:Luciferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D010449:Peptide Mapping; D011489:Protein Denaturation; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors; D014357:Trypsin
C091732:BCL2-associated athanogene 1 protein; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; C494371:HSPA8 protein, human; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D008156:Luciferases; D014357:Trypsin; D000251:Adenosine Triphosphatases