title: BiP mutants that are unable to interact with endoplasmic reticulum DnaJ proteins provide insights into interdomain interactions in BiP.
abstract: The heat shock protein (Hsp)70 family of molecular chaperones interacts with unfolded proteins through a C-terminal substrate-binding domain (SBD) that is controlled by nucleotide binding to the N-terminal domain. The ATPase cycle is regulated by cochaperones, including DnaJ proteins that accelerate ATP hydrolysis to stabilize the Hsp70-substrate complex. We found that R197 in hamster BiP, which resides at the surface of the nucleotide-binding domain, is critical for both association with endoplasmic reticulum DnaJ proteins and interaction with the SBD. Decreasing the positive charge at this residue enhanced basal ATPase activity, destabilized interaction with the SBD, and reduced substrate release both in vitro and in vivo. Mutation of three glutamic acids in the SBD mimicked many of these effects. Our data provide insights into communications between the two domains and suggest a mechanism by which DnaJ proteins increase ATP hydrolysis.
authors.affiliation: Department of Genetics and Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Walid
authors.initials: W
authors.lastname: Awad
authors.forename: Isaac
authors.initials: I
authors.lastname: Estrada
authors.forename: Ying
authors.initials: Y
authors.lastname: Shen
authors.forename: Linda M
authors.initials: LM
authors.lastname: Hendershot
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019943:Amino Acid Substitution / Q000235:genetics*; D000818:Animals; D001120:Arginine / Q000737:chemistry / Q000235:genetics; D019556:COS Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D050956:HSP40 Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D017386:Sequence Homology, Amino Acid
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D050956:HSP40 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D001120:Arginine; D000251:Adenosine Triphosphatases