title: Proteolysis prevents in vivo chimeric fusion protein import into yeast mitochondria. Cytosolic cleavage and subcellular distribution.
abstract: The in vivo import of liver mitochondrial aldehyde dehydrogenase was investigated in yeast by constructing fusion proteins between its leader sequence and beta-galactosidase. Only 7% of the protein was imported. If 21 or 71 amino acids from the mature portion of aldehyde dehydrogenase were included in the construct, 40% was imported. The protein remaining in cytosol was sequenced. When the leader was fused directly to beta-galactosidase, the first 7 residues of the leader were missing. When 21 residues of mature aldehyde dehydrogenase were included, the entire leader plus 6 residues of the mature portion were missing; if 71 residues of mature aldehyde dehydrogenase were included, the first residue found corresponds to the 66th residue of the mature portion. When the leader was fused directly to beta-galactosidase, no processing of the imported protein occurred, and the N-terminal amino acid was blocked, presumably by acetylation. If the 21-amino acid insert was included, processing occurred. A modified leader sequence lacking the three-amino acid linker (RGP) was imported but not processed, just as we found in vitro (Thornton, K., Wang, Y., Weiner, H., and Gorenstein, D.G. (1993) J. Biol. Chem. 268, 19906-19914). The less than 100% import of pre-aldehyde dehydrogenase was due to the action of a post-translational protease attack which prevented import by destroying the leader peptide segment.
authors.affiliation: Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907-1153, USA.
authors.forename: J
authors.initials: J
authors.lastname: Zhou
authors.forename: Y
authors.initials: Y
authors.lastname: Bai
authors.forename: H
authors.initials: H
authors.lastname: Weiner
mesh_terms: D000444:Aldehyde Dehydrogenase / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003600:Cytosol / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D011993:Recombinant Fusion Proteins; D000444:Aldehyde Dehydrogenase; D001616:beta-Galactosidase