Functional divergence between co-chaperones of Hsc70.
The ATPase cycle of the chaperone Hsc70 is regulated by co-chaperones; Hsp40/DnaJ-related proteins stimulate ATP hydrolysis by Hsc70 and can bind unfolded polypeptides themselves. Conversely, various nucleotide exchange factors (NEFs) stimulate ADP-ATP exchange by Hsc70. We analyzed the purified Hsp40-related co-chaperones DJA1 (Hdj2) and DJA2 (Hdj3) and found that they had a distinct pattern of binding to a range of polypeptides. DJA2 alone could stimulate Hsc70-mediated refolding of luciferase in the absence of NEF, whereas DJA1 was much less active. The addition of the Bag1 NEF increased refolding by Hsc70 and DJA2, as did the newly characterized NEF Hsp110, but each NEF had a different optimal concentration ratio to Hsc70. Notably, the NEF HspBP1 could not increase refolding by Hsc70 and DJA2 at any concentration, and none of the NEFs improved the refolding activity with DJA1. Instead, DJA1 was inhibitory of refolding with DJA2 and Hsc70. All combinations of DJA1 or DJA2 with the three NEFs stimulated the Hsc70 ATPase rate, although Hsp110 became less effective with increasing concentrations. A chimeric DJA2 having its Hsc70-stimulatory J domain replaced with that of DJA1 was functional for polypeptide binding and ATPase stimulation of Hsc70. However, it could not support efficient Hsc70-mediated refolding and also inhibited refolding with DJA2 and Hsc70. These results suggest a more complex model of Hsc70 mechanism than has been previously thought, with notable functional divergence between Hsc70 co-chaperones.
Tzankov
Stefan
S
Department of Biochemistry, McGill University, Montreal, Quebec H3G 1Y6, Canada.
Wong
Michael J H
MJ
Shi
Kun
K
Nassif
Christina
C
Young
Jason C
JC
D000255:Adenosine Triphosphate / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002417:Cattle; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005326:Fetal Proteins / Q000235:genetics / Q000378:metabolism*; D050883:HSC70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D050957:HSP110 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D050956:HSP40 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D051379:Mice; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D017510:Protein Folding; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
C091732:BCL2-associated athanogene 1 protein; D004268:DNA-Binding Proteins; C107481:DNAJA1 protein, human; C494647:DNAJB1 protein, human; C476535:DNAJC14 protein, human; D005326:Fetal Proteins; D050883:HSC70 Heat-Shock Proteins; D050957:HSP110 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; C494371:HSPA8 protein, human; D018832:Molecular Chaperones; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D000255:Adenosine Triphosphate