N-terminal tail export from the mitochondrial matrix. Adherence to the prokaryotic "positive-inside" rule of membrane protein topology.
Export of N-terminal tails of mitochondrial inner membrane proteins from the mitochondrial matrix is a membrane potential-dependent process, mediated by the Oxa1p translocation machinery. The hydrophilic segments of these membrane proteins, which undergo export, display a characteristic charge profile where intermembrane space-localized segments bear a net negative charge, whereas those remaining in the matrix have a net positive one. Using a model protein, preSu9(1-112)-dihydrofolate reductase (DHFR), which undergoes Oxa1p-mediated N-tail export, we demonstrate here that the net charge of N- and C-flanking regions of the transmembrane domain play a critical role in determining the orientation of the insertion process. The N-tail must bear a net negative charge to be exported to the intermembrane space. Furthermore, a net positive charge of the C-terminal region supports this N-tail export event. These data provide experimental evidence that protein export in mitochondria adheres to the "positive-inside" rule, described for sec-independent sorting of membrane proteins in prokaryotes. We propose here that the importance of a charge profile reflects a need for specific protein-protein interactions to occur in the export reaction, presumably at the level of the Oxa1p export machinery.
Institut für Physiologische Chemie der Universität München, Goethestrasse 33, 80336 München, Germany.
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Rojo
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Guiard
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Neupert
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Stuart
D001692:Biological Transport; D002467:Cell Nucleus / Q000378:metabolism; D003576:Electron Transport Complex IV; D008565:Membrane Proteins / Q000737:chemistry; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins; D016296:Mutagenesis; D009492:Neurospora crassa / Q000201:enzymology; D009687:Nuclear Proteins / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae; D055672:Static Electricity; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry / Q000235:genetics
D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D009687:Nuclear Proteins; C090858:OXA1 protein; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D013762:Tetrahydrofolate Dehydrogenase; D003576:Electron Transport Complex IV; D006180:Proton-Translocating ATPases