Skip to main content
SpringerLink
Log in

Functional properties of purified and reconstituted mitochondrial metabolite carriers

  • Published:
Journal of Bioenergetics and Biomembranes Aims and scope Submit manuscript

Abstract

Eight mitochondrial carrier proteins were solubilized and purified in the authors' laboratories using variations of a general procedure based on hydroxyapatite and Celite chromatography. The molecular mass of all the carriers ranges between 28 and 34 kDa on SDS-PAGE. The purified carrier proteins were reconstituted into liposomes mainly by using a method of detergent removal by hydrophobic chromatography on polystyrene beads. The various carriers were identified in the reconstituted state by their kinetic properties. A complete set of basic kinetic data including substrate specificity, affinity, interaction with inhibitors, and activation energy was obtained. These data closely resemble those of intact mitochondria, as far as they are available from the intact organelle. Mainly on the basis of kinetic data, the asymmetric orientation of most of the reconstituted carrier proteins were established. Several of their functional properties are significantly affected by the type of phospholipids used for reconstitution. All carriers which have been investigated in proteoliposomes function according to a simultaneous (sequential) mechanism of transport; i.e., a ternary complex, made up of two substrates and the carrier protein, is involved in the catalytic cycle. The only exception was the carnitine carrier, where a ping-pong mechanism of transport was found. By reaction of particular cysteine residues with mercurial reagents, several carriers could be reversibly converted to a functional state different from the various physiological transport modes. This “unphysiological” transport mode is characterized by a combination of channel-type and carrier-type properties.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Aquila, H. Link, T. A., and Klingenberg, M. (1987).FEBS Lett. 212, 1–9.

    PubMed  Google Scholar 

  • Bisaccia, F., and Palmieri, F. (1984).Biochim. Biophys. Acta 766, 386–394.

    PubMed  Google Scholar 

  • Bisaccia, F., Indiveri, C., and Palmieri, F. (1985).Biochim. Biophys. Acta 810, 362–369.

    Google Scholar 

  • Bisaccia, F., Indiveri, C., and Palmieri, F. (1988).Biochim. Biophys. Acta 933, 229–240.

    PubMed  Google Scholar 

  • Bisaccia, F., De Palma, A., and Palmieri, F. (1989).Biochim. Biophys. Acta 977, 171–176.

    PubMed  Google Scholar 

  • Bisaccia, F., De Palma, A., Prezioso, G., and Palmieri, F. (1990).Biochim. Biophys. Acta 1019, 250–256.

    PubMed  Google Scholar 

  • Bisaccia, F., De Palma, A., and Palmieri, F. (1992)Biochim. Biophys. Acta 1106, 291–296.

    PubMed  Google Scholar 

  • Bisaccia, F., De Palma, A., Dierks, T., Krämer, R., and Palmieri, F. (1993).Biochim. Biophys. Acta 1142, 139–145.

    PubMed  Google Scholar 

  • Bolli, R., Nalecz, K. A., and Azzi, A. (1989).J. Biol. Chem. 264, 18024–18030.

    PubMed  Google Scholar 

  • Claeys, D., and Azzi, A. (1989).J. Biol. Chem. 264, 14627–14630.

    PubMed  Google Scholar 

  • Cleland, W. W. (1970). InThe Enzymes, Vol. 2 (Boyer, P. D., ed.), Academic Press, New York, pp. 1–65.

    Google Scholar 

  • Crompton, M., Palmieri, F., Capano, M., and Quagliariello, E. (1975).Biochem. J. 146, 667–673.

    PubMed  Google Scholar 

  • de Kruiff, B. (1985). InProgress in Protein-Lipid Interaction, Vol. I (Watts, A., and de Pont, J. J. H. H. M., eds.), Elsevier, Amsterdam.

    Google Scholar 

  • De Pinto, V., Tommasino, M., Palmieri, F., and Kadenbach, B. (1982).FEBS Lett. 148, 103–106.

    PubMed  Google Scholar 

  • Dierks, T., and Krämer, R. (1988).Biochim. Biophys. Acta 937, 112–126.

    PubMed  Google Scholar 

  • Dierks, T., Riemer, E., and Krämer, R. (1988).Biochim. Biophys. Acta 943, 231–244.

    PubMed  Google Scholar 

  • Dierks, T., Salentin, A., Heberger, C., and Krämer, R. (1990a).Biochim. Biophys. Acta 1028, 268–280.

    PubMed  Google Scholar 

  • Dierks, T., Salentin, A., and Krämer, R. (1990b).Biochim. Biophys. Acta 1028, 281–288.

    PubMed  Google Scholar 

  • Duykaerts, C., Sluse-Goffart, C., Fux, J. P., Sluse, F. E., and Liebecq, C. (1980).Eur. J. Biochem. 106, 1–6.

    PubMed  Google Scholar 

  • Indiveri, C., Palmieri, F., Bisaccia, F., and Krämer, R. (1987a).Biochim. Biophys. Acta 890, 310–318.

    PubMed  Google Scholar 

  • Indiveri, C., Krämer, R., and Palmieri, F. (1987b).J. Biol. Chem. 262, 15979–15984.

    PubMed  Google Scholar 

  • Indiveri, C., Capobianco, L., Krämer, R., and Palmieri, F. (1989a).Biochim. Biophys. Acta 977, 187–193.

    PubMed  Google Scholar 

  • Indiveri, C., Dierks, T., Krämer, R., Palmieri, F. (1989b).Biochim. Biophys. Acta 977, 194–199.

    PubMed  Google Scholar 

  • Indiveri, C., Tonazzi, A., and Palmieri, F. (1990).Biochim. Biophys. Acta 1020, 81–86.

    PubMed  Google Scholar 

  • Indiveri, C., Dierks, T., Krämer, R., and Palmieri, F. (1991a).Eur. J. Biochem. 198, 339–347.

    PubMed  Google Scholar 

  • Indiveri, C., Tonazzi, a., Prezioso, G., and Palmieri, F. (1991b).Biochim. Biophys. Acta 1065, 231–238.

    PubMed  Google Scholar 

  • Indiveri, C., Tonazzi, A., and Palmieri, F. (1991c).Biochim. Biophys. Acta 1069, 110–116.

    PubMed  Google Scholar 

  • Indiveri, C., Tonazzi, A., and Palmieri, F. (1992a).Eur. J. Biochem. 207, 449–454.

    PubMed  Google Scholar 

  • Indiveri, C., Tonazzi, A., Dierks, T., Krämer, R., and Palmieri, F. (1992b).Biochim. Biophys. Acta 1140, 53–58.

    PubMed  Google Scholar 

  • Indiveri, C., Prezioso, G., Dierks, T., Krämer, R., and Palmieri, F. (1993a).Biochim. Biophys. Acta,1143, 310–318.

    PubMed  Google Scholar 

  • Indiveri, C., Tonazzi, A., and Palmieri, F. (1993b). Submitted.

  • Kadenbach, B., Mende, P., Kolbe, H. V. J., Stipani, I., and Palmieri, F. (1982).FEBS Lett. 139, 109–112.

    PubMed  Google Scholar 

  • Kaplan, R. S., Pratt, R. D., and Pedersen, P. L. (1986).J. Biol. Chem. 261, 12767–12773.

    PubMed  Google Scholar 

  • Klingenberg, M. (1981).Nature 290, 449–454.

    PubMed  Google Scholar 

  • Klingenberg, M. (1989).Arch. Biochem. Biophys. 270, 1–14.

    PubMed  Google Scholar 

  • Kolbe, H. V. J., Böttrich, J., Genchi, G., Palmieri, F., and Kadenbach, B. (1981).FEBS Lett. 124, 265–269.

    PubMed  Google Scholar 

  • Kolbe, H. V. J., Costello, D., Wong, A., Lu, R. C., and Wohlrab, H., J. (1984).J. Biol. Chem. 259, 9115–9120.

    PubMed  Google Scholar 

  • Krämer, R. (1982).Biochim. Biophys. Acta 693, 269–304.

    Google Scholar 

  • Krämer, R. (1983).Biochim. Biophys. Acta 735, 145–159.

    PubMed  Google Scholar 

  • Krämer, R., and Heberger, C. (1986).Biochim. Biophys. Acta 863, 289–296.

    PubMed  Google Scholar 

  • Krämer, R., and Klingenberg, M. (1977).FEBS Lett. 82, 363–367.

    PubMed  Google Scholar 

  • Krämer, R., and Klingenberg, M. (1979).Biochemistry 18, 209–4215.

    Google Scholar 

  • Krämer, R., and Klingenberg, M. (1980a)Biochemistry 19, 56–560.

    Google Scholar 

  • Krämer, R., and Klingenberg, M. (1980b).FEBS Lett. 119, 257–260.

    PubMed  Google Scholar 

  • Krämer, R., and Klingenberg, M. (1982).Biochemistry 21, 082–1089.

    Google Scholar 

  • Krämer, R., and Palmieri, F. (1989).Biochim. Biophys. Acta 974, 1–23.

    PubMed  Google Scholar 

  • Krämer, R., and Palmieri, F. (1992). InMolecular Mechanisms in Bioenergetics (Ernster, L., ed.), Elsevier, Amsterdam, pp. 359–384.

    Google Scholar 

  • Krämer, R., Kurzinger, G., and Heberger, C. (1986).Arch. Biochem. Biophys. 251, 166–174.

    PubMed  Google Scholar 

  • Lin, C. S., and Klingenberg, M. (1980).FEBS Lett. 113, 299–303.

    PubMed  Google Scholar 

  • Meisner, H., Palmieri, F., and Quagliariello, E. (1972).Biochemistry 11, 949–955.

    PubMed  Google Scholar 

  • Mende, P., Kolbe, H. V. J., Kadenbach, B., Stipani, I., and Palmieri, F. (1982).Eur. J. Biochem. 128, 91–95.

    PubMed  Google Scholar 

  • Palmieri, F., Prezioso, G., Quagliariello, E., and Klingenberg, M. (1971).Eur. J. Biochem. 22, 66–74.

    PubMed  Google Scholar 

  • Palmieri, F., Quagliariello, E., and Klingenberg, M. (1972a).Eur. J. Biochem. 29, 408–416.

    PubMed  Google Scholar 

  • Palmieri, F., Stipani, I., Quagliariello, E., and Klingenberg, M. (1972b).Eur. J. Biochem. 26, 587–594.

    PubMed  Google Scholar 

  • Palmieri, F., Bisaccia, F., Capobianco, L., Iacobazzi, V., Indiveri, C., and Zara, V. (1990).Biochim. Biophys. Acta 1081, 147–150.

    Google Scholar 

  • Palmieri, F., Bisaccia, F., Capobianco, L., Dolce, V., Fiermonte, G., Iacobazzi, V., and Zara, V., (1993).J. Bioenerg. Biomembr.,25, 493–501.

    PubMed  Google Scholar 

  • Quagliariello, E., Passarella, S., and Palmieri, F. (1974). InDynamics of energy-Transducing Membranes (Ernster, L., Estabrook, A., and Slater, E. C., eds.), Elsevier, Amsterdam, pp. 483–495.

    Google Scholar 

  • Riccio, P., Aquila, H., and Klingenberg, M. (1975).FEBS Lett. 56, 133–138.

    PubMed  Google Scholar 

  • Runswick, M. J., Powell, S. J., Nyren, P., Walker, J. E. (1987).EMBO J. 6, 1367–1373.

    PubMed  Google Scholar 

  • Runswick, M. J., Walker, J. E., Bisaccia, F., Iacobazzi, V., and Palmieri, F. (1990).Biochemistry 29, 11033–11040.

    PubMed  Google Scholar 

  • Sluse, F. E., Duyckaerts, C., Liebecq, S., and Sluse-Goffart, M. (1979).Eur. J. Biochem. 100, 3–17.

    PubMed  Google Scholar 

  • Sluse, F. E., Evens, A., Dierks, T., Duyckaerts, C., Sluse-Goffart, C. M., and Krämer, R. (1991).Biochim. Biophys. Acta 1058, 329–338.

    PubMed  Google Scholar 

  • Stappen, R., and Krämer, R. (1993a).Biochim. Biophys. Acta, in press.

  • Stappen, R., and Krämer, R. (1993b). submitted.

  • Stipani, I., and Palmieri, F. (1983).FEBS Lett. 161, 269–274.

    PubMed  Google Scholar 

  • Ueno, M., Tanford, C., and Reynolds, J. A. (1984).Biochemistry 23, 3070–3076.

    PubMed  Google Scholar 

  • Walker, J. E., and Runswick, M. J. (1993).J. Bioenerg. Biomembr.,25, 435–446.

    PubMed  Google Scholar 

  • Wohlrab, H. (1980).J. Biol. Chem. 255, 8170–8173.

    PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Pharmaco-Biology, Laboratory of Biochemistry and Molecular Biology, University of Bari, Bari, Italy

    F. Palmieri, C. Indiveri & F. Bisaccia

  2. Institut für Biotechnologie I, Forschungszentrum Jülich, Jülich, Germany

    R. Krämer

Authors
  1. F. Palmieri
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. C. Indiveri
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. F. Bisaccia
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. R. Krämer
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Palmieri, F., Indiveri, C., Bisaccia, F. et al. Functional properties of purified and reconstituted mitochondrial metabolite carriers. J Bioenerg Biomembr 25, 525–535 (1993). https://doi.org/10.1007/BF01108409

Download citation

  • Received:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF01108409

Key words

Search

Navigation