Mitochondrial substrate carriers.
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Amino Acid Transport Systems, Acidic↗
Biomedical subjects
V Zara
Publications and source records attributed to V Zara.
The cleavable presequence is not essential for import and assembly of the phosphate carrier of mammalian mitochondria but enhances the specificity and efficiency of import.
The phosphate carrier (PiC) of mammalian mitochondria is synthesized with a cleavable presequence, in contrast to other members of the mitochondrial family of inner membrane carrier proteins. The precursor of PiC is efficiently imported, proteolytically processed, and correctly assembled in isolated mitochondria. Here we report that a presequence-deficient PiC was imported with an efficiency of about 50% as compared with the authentic precursor of PiC. This mature-sized PiC was correctly assembled, demonstrating that the presequence is not essential for the assembly pathway. We found the following functions for the PiC presequence. (i) The presequence by itself was able to target a passenger protein to mitochondria with a low efficiency, suggesting that the mammalian PiC contains multiple targeting signals, the more efficient one(s) present in the mature protein part. (ii) Deletion of the presequence allowed a more efficient heterologous import of mammalian PiC into mitochondria from Saccharomyces cerevisiae and Neurospora crassa, indicating an important role of the presequence in determining the specificity of PiC import. (iii) Import of the presequence-deficient PiC required a higher membrane potential across the inner membrane than that of the presequence-carrying form. Therefore, the presequence also enhances the translocation of PiC into the inner membrane.
Animals↗
Biogenesis of the mitochondrial phosphate carrier.
The mitochondrial phosphate carrier (PiC) is a member of the family of inner-membrane carrier proteins which are generally synthesized without a cleavable presequence. Surprisingly, the cDNA sequences of bovine and rat PiC suggested the existence of an amino-terminal extension sequence in the precursor of PiC. By expressing PiC in vitro, we found that PiC is indeed synthesized as a larger precursor. This precursor was imported and proteolytically processed by mitochondria, whereby the correct amino-terminus of the mature protein was generated. Import of PiC showed the characteristics of mitochondrial protein uptake, such as dependence on ATP and a membrane potential and involvement of contact sites between mitochondrial outer and inner membranes. The precursor imported in vitro was correctly assembled into the functional form, demonstrating that the authentic import and assembly pathway of PiC was reconstituted when starting with the presequence-carrying precursor. These results are discussed in connection with the recently postulated role of PiC as an import receptor located in the outer membrane.
Amino Acid Sequence↗
Characterization of pore-forming activity in liver mitochondria from Anguilla anguilla. Two porins in mitochondria?
A fast purification procedure for the isolation and purification of eukaryotic porin (De Pinto et al., (1987) Biochim. Biophys. Acta 905, 499-502) was applied to liver mitochondria of the fish Anguilla anguilla. A protein preparation was obtained which formed slightly anionically selective pores in reconstitution experiments with lipid bilayer membranes. The distribution of single-channel conductances had two maxima of 2.4 nS and 4.0 nS in 1 M KCl. Sodium dodecylsulfate electrophoretograms of the protein preparation showed the presence of two bands of very similar electrophoretic mobility (32 and 32.5 kDa). Both bands cross-reacted with antibodies raised against purified bovine heart porin and with antibodies raised against the 19 amino acids N-terminal end of human porin. No cross-reactivity was observed with antibodies against yeast porin. The peptide maps of the two bands showed slight differences. The possibility of the presence of two different porins in liver mitochondria of Anguilla anguilla is discussed. An extensive immunological comparison of different mitochondrial porins is presented.
Anguilla↗
Structural and functional properties of mitochondrial anion carriers.
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Animals↗
Immunological characterization of the mitochondrial 2-oxoglutarate carrier from liver and heart. Organ specificity.
Antibodies have been prepared against the 2-oxoglutarate transport proteins purified from bovine heart and rat liver mitochondria. The anti-heart antiserum cross-reacts with the 2-oxoglutarate carrier (OGC) from beef, pig, rat and rabbit heart, but not with the OGC from liver of the same animals. Conversely, the anti-liver antiserum recognizes the carrier protein from liver of all species tested but not from heart. Immunoinactivation of oxoglutarate transport activity by the antibodies is also tissue specific. Peptide maps of purified OGC show structural differences between the carrier from heart and liver of the same animal species. These results indicate the existence of isoforms of the OGC in heart and liver.
Animals↗
Effect of anthracycline antibiotics on the reconstituted mitochondrial tricarboxylate carrier.
The effect of anthracycline antibiotics on the activity of the partially purified and reconstituted tricarboxylate carrier system of the rat liver mitochondria was studied. It was found that the citrate/citrate exchange activity is inhibited by Br-daunomycin and with less potency by doxorubicin, daunomycin, epirubicin and idarubicin. The inhibition of the citrate transport activity is concentration and time-dependent. Cardiolipin protects against the inhibition by Br-daunomycin and the reconstituted citrate transport activity depends upon the ratio of cardiolipin/Br-daunomycin.
Animals↗
Inhibition and labelling of the mitochondrial 2-oxoglutarate carrier by eosin-5-maleimide.
Unlike hydrophobic maleimides, eosin-5-maleimide and to a lesser extent other relatively polar maleimides inhibit the 2-oxoglutarate carrier of bovine heart mitochondria. The impermeable eosin-5-maleimide labels the 2-oxoglutarate carrier in intact mitochondria but not in submitochondrial particles. 2-Oxoglutarate protects the carrier against inactivation by eosin-5-maleimide and decreases the fluorescence associated with the purified protein. Other anions which are not substrates of the carrier have no protective effect. It is concluded that sulfhydryl groups essential for the activity of the 2-oxoglutarate carrier are located at the cytosolic face of the inner mitochondrial membrane. They appear to be present at the substrate-binding site and located in a hydrophilic environment.
Biological Transport↗