Biogenesis of ISP6, a small carboxyl-terminal anchored protein of the receptor complex of the mitochondrial outer membrane.
To study the biogenesis of ISP6, an outer membrane component of the mitochondrial protein translocation complex, two fusion proteins have been made by fusing ISP6 to either the carboxyl- or amino-terminal end of the mouse dihydrofolate reductase (DHFR). In vitro import experiments showed that when DHFR was placed at the carboxyl-terminal end of ISP6, the resulting fusion protein 6-DHFR inserted into mitochondrial membrane less efficiently than the other form of the fusion proteins. In vivo this fusion protein lost its ability to suppress the temperature-sensitive phenotype of an isp42 mutant, while the other fusion protein DHFR-6, which was found targeted correctly to mitochondria, suppressed the mutant as well as the wild-type ISP6. Further analysis showed that the binding and insertion of DHFR-6 to mitochondrial outer membrane was not affected by deletion of either of the two mitochondrial protein receptors or by the predigestion of mitochondrial surface proteins prior to import. Additional data indicated that ISP42, which closely associates with ISP6 in the translocation complex, does not likely play the role of a targeting partner for ISP6. In summary, these data suggest that ISP6 may target to mitochondria by sequences at its carboxyl terminus and that the import process of ISP6 is most likely distinct from that of most other mitochondrial precursors, which are recognized by protein receptors on mitochondrial surface.
Cao
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Department of Biochemistry and Biophysics, University of North Carolina Medical School, Chapel Hill 27599-7260.
Douglas
M G
MG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003600:Cytosol / Q000378:metabolism; D017931:DNA Primers; D005656:Fungal Proteins / Q000096:biosynthesis*; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013489:Suppression, Genetic; D013762:Tetrahydrofolate Dehydrogenase / Q000096:biosynthesis; D014158:Transcription, Genetic
D017931:DNA Primers; D005656:Fungal Proteins; D008565:Membrane Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C082532:TOM6 protein, S cerevisiae; D013762:Tetrahydrofolate Dehydrogenase