Proteins of the Hsp70 family of ATPases interact with a conserved domain of their J-protein partners, the J-domain, to function in numerous cellular processes. We have studied the interaction of BiP, an Hsp70 family member in the lumen of the endoplasmic reticulum, with the J-domain of Sec63p, a component of the Sec complex involved in post-translational protein translocation across the endoplasmic reticulum membrane. In a real-time solid phase binding assay, BiP binds to the immobilized Sec complex or to a fusion protein of the J-domain and glutathione S-transferase in a reaction that requires ATP hydrolysis. In the final complex, BiP is bound in the ADP form with its peptide binding pocket occupied. An intact peptide binding pocket is required for this interaction. Our experiments suggest that the activation of BiP by the J-domain involves a transient contact between these components, and that in the absence of physiological substrates, J-activated BiP binds even to the J-proteins themselves.

• PMID: 10400622
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Journal Article | Research Support, U.S. Gov't, P.H.S.

Authors

Misselwitz B, Staeck O, Matlack KE, Rapoport TA

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