A precursor protein partly translocated into yeast mitochondria is bound to a 70 kd mitochondrial stress protein.
We have probed the environment of a precursor protein stuck in mitochondrial import sites using cleavable bifunctional crosslinking reagents. The stuck precursor was crosslinked to a 70 kd protein which, by immunological techniques, was shown to be a matrix protein. The protein was purified to homogeneity by ATP-Sepharose chromatography and partially sequenced. Fourteen of its 15 N-terminal amino acids were identical to residues 24-38 of the protein encoded by the nuclear gene SSC1, which had been proposed to encode a dnaK-like 70 kd mitochondrial stress protein. Our data imply that this mitochondrial hsp70 is made with a cleavable matrix-targeting sequence composed of 23 residues. The complex containing stuck precursor, mitochondrial hsp70, and ISP42 could be solubilized from mitochondria by the non-ionic detergent Triton X-100 even without crosslinking, suggesting tight association of these three components. As the stuck precursor is arrested at an early stage of translocation, mitochondrial hsp70 may initiate the events that lead to refolding of imported precursors in the matrix space.
Scherer
P E
PE
Biocenter, University of Basel, Switzerland.
Krieg
U C
UC
Hwang
S T
ST
Vestweber
D
D
Schatz
G
G
D001692:Biological Transport; D002678:Chimera; D003432:Cross-Linking Reagents / Q000378:metabolism; D005656:Fungal Proteins / Q000276:immunology*; D006360:Heat-Shock Proteins / Q000276:immunology / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins; D016253:Microscopy, Immunoelectron; D008928:Mitochondria / Q000378:metabolism* / Q000648:ultrastructure; D033681:Mitochondrial Membrane Transport Proteins; D011498:Protein Precursors / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins
D003432:Cross-Linking Reagents; D005656:Fungal Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D033681:Mitochondrial Membrane Transport Proteins; D011498:Protein Precursors; D029701:Saccharomyces cerevisiae Proteins; C066555:Tom40 protein, S cerevisiae