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 ars enal of cochaperones, including the anti-apoptotic Bag-IM protein, which re gulate the activities of Hsp70s by controlling their ATPase cycles. To eluc idate the regulatory function of Bag-1M, we determined its influence on nuc leotide 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 a s they stimulated the ADP dissociation rate of Hsc70 and Hsp70 up to 900-fo ld. The N-terminal domain of Bag-IM decreased the affinity of Bag-IM for Hs c70/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-depen dent refolding of luciferase in the absence of P-i. Surprisingly, under phy siological conditions, i.e. low Bag-IM concentrations and presence of P-i, Bag-IM activates the chaperone action of Hsc70/Hsp70 in luciferase refoldin g. Bag-IM accelerated ATP-triggered substrate release by Hsc70/Hsp70. We pr opose that Bag-IM acts as substrate discharging factor for Hsc70 and Hsp70.