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  • 1
    ISSN: 1573-6881
    Keywords: Mitochondria ; transport ; overexpression ; dicarboxylate carrier ; ACR1 gene ; succinate-fumarate exchange ; ARG11 gene; ornithine carrier ; arginine biosynthesis ; yeast ; metabolism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract The genome of Saccharomyces cerevisiae encodes 35 members of a family proteins thattransport metabolites and substrates across the inner membranes of mitochondria. They includethree isoforms of the ADP/ATP translocase and the phosphate and citrate carriers. At the startof our work, the functions of the remaining 30 members of the family were unknown. We areattempting to identify these 30 proteins by overexpression of the proteins in specially selectedhost strains of Escherichia coli that allow the carriers to accumulate at high levels in the formof inclusion bodies. The purified proteins are then reconstituted into proteoliposomes wheretheir transport properties are studied. Thus far, we have identified the dicarboxylate,succinate-fumarate and ornithine carriers. Bacterial overexpression and functional identification, togetherwith characterization of yeast knockout strains, has brought insight into the physiologicalsignificance of these transporters. The yeast dicarboxylate carrier sequence has been used toidentify the orthologous protein in Caenorhabditis elegans and, in turn, this latter sequencehas been used to establish the sequence of the human ortholog.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-6881
    Keywords: Mitochondrial porin ; VDAC ; Zea mays, Pisum sativum ; sterol ; reconstitution ; voltage dependence ; lipid bilayer membrane
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Water-soluble porins were prepared from native mitochondrial porins isolated from different plants (pea and corn). In the water-soluble form the porins have lost their channel-forming properties. The water-soluble porins were investigated for the influence of different sterols on their membrane activity and their channel-forming properties in lipid bilayer membranes. Our experiments demonstrated that the water-soluble porins regained channel forming activity when the protein was preincubated with different sterols in the presence of a detergent. The channels formed in lipid bilayer membranes after this procedure regain in many but not all cases the original properties of the native mitochondrial porins. Preincubation with other sterols led to a change in the single-channel conductance or to a complete loss of the voltage dependence. The sterols had also a strong influence on the channel-forming activity of the porins. Preincubation of water-soluble pea porin with the plant sterol β-sitosterol resulted in a considerable higher channel-forming activity than with all the other sterols used for preincubation. The role of the sterols in the channel-forming complex is discussed.
    Type of Medium: Electronic Resource
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