Mitochondrial Hsp70 cannot replace BiP in driving protein translocation into the yeast endoplasmic reticulum.
To determine whether mitochondrial hsp70 (mHsp70) could substitute for the endoplasmic retuculum (ER) Hsp70 (BiP) during protein translocation, we assembled ER-derived reconstituted proteoliposomes supplemented with either protein. We found that only BiP restored translocation in kar2 mutant vesicles and stimulated translocation approximately 3-fold in wild type proteoliposomes. mHsp70 associated poorly with both a BiP binding (DnaJ) domain of Sec63p and an ER precursor, and its ATPase activity was poorly enhanced upon incubation with the DnaJ domain. In contrast, BiP bound to the Sec63p-DnaJ domain in an ATP-dependent manner and its ATPase activity was stimulated significantly by this polypeptide. We conclude that mHsp70 is unable to support protein translocation into the ER because it fails to associate productively with Sec63p and a precursor.
Brodsky
J L
JL
Department of Biological Sciences, University of Pittsburgh, PA 15260, USA. jbrodsky+@pitt.edu
Bäuerle
M
M
Horst
M
M
McClellan
A J
AJ
D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005656:Fungal Proteins / Q000096:biosynthesis*; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis*; D008928:Mitochondria / Q000378:metabolism*; D014176:Protein Biosynthesis; D012441:Saccharomyces cerevisiae / Q000378:metabolism* / Q000648:ultrastructure
D005656:Fungal Proteins; D018840:HSP70 Heat-Shock Proteins; C060153:KAR2 protein, yeast