Abstract
Genes encoding VDAC proteins containing specific site-directed amino acid alterations were introduced into wild-typeSaccharomyces cerevisiae. The mutant VDAC proteins form channels with ion selectivities very different from that of the wild-type channel. Therefore, the resulting yeast strains express two different genes capable of coding for functional, yet distinct, VDAC channels. If VDAC were an oligomeric channel, analysis of VDAC from these strains should have revealed not only the presence of channels with wild-type or mutant selectivity but also channels with intermediate selectivities. While channels with wild-type and mutant selectivities were observed with approximately equal frequency, no channels with intermediate selectivity were observed. Sufficient observations were performed with two different mutant genes (K61E.K65E and K19E.K61E) that the likelihood of having missed hybrid channels was less than 1 in 107. These findings favor the hypothesis that each functional VDAC channel is composed of a single 30-kDa polypeptide chain.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Blachly-Dyson, E., Peng, S. Colombini, M., and Forte, M. (1990). Selectivity Changes in Site-directed Mutants of the VDAC Ion Channel: Structural Implications,Science 247 1233–1236.
Colombini, M. (1980). Structure and Model of a Voltage-Dependent Anion-Selective Channel (VDAC) Located in the Outer Mitochondrial Membrane,Ann. NY. Acad. Sci. 341 552–563.
Colombini, M. (1987). Characterization of Channels Isolated from Plant Mitochondria,Methods Enzymol. 48 465–475.
Colombini, M. (1989). Voltage Gating in the Mitochondrial Channel, VDAC,J. Membr. Biol. 111 103–111.
Colombini, M., Peng, S., Blachly-Dyson, E., and Forte, M. (1991). Probing the Molecular and Structural Changes of a Voltage-Gated Channel,Methods Enzymol. 207, in press.
Forte, M., Guy, R., and Mannella, C. (1987). Molecular Genetics of the VDAC Ion Channel: Structural Model and Sequence Analysis,J. Bioenerg. Biomembr. 19 341–350.
Holden, M., and Colombini, M. (1988). The Mitochondrial Outer Membrane Channel, VDAC, Is Modulated by a Soluble Protein,FEBS Lett. 241 105–109.
Linden, M., and Gellerfors, P. (1983). Hydrodynamic Properties of Porin Isolated from Outer Membrane Of Rat Liver Mitochondria,Biochim. Biophys. Acta 736 125–129.
Mangan, P. S., and Colombini, M. (1987). Ultrasteep Voltage Dependence in a Membrane Channel,Proc. Natl. Acad. Sci. USA 84 4896–4900.
Mannella, C. (1986). Mitochondrial Outer Membrane Channel (VDAC, Porin) Two-Dimensional Crystals fromNeurospora, Methods Enzymol. 125 595–610.
Mannella, C. (1987). Electron Microscopy and Image Analysis of the Mitochondrial Outer Membrane Channel, VDAC,J. Bioenerg. Biomembr. 19 329–340.
Montal, M., and Mueller, P. (1972). Formation of Bimolecular Membranes from Lipid Monolayers and a Study of Their Electrical Properties,Proc. Natl. Acad. Sci. USA 69 3561–3566.
Roos, N., Benz, R., and Bridczka, D. (1982). Identification and Characterization of the Pore-Forming Protein in the Outer Membrane of Rat Liver Mitochondria,Biochim. Biophys. Acta 686 204–214.
Schein, S. J., Colombini, M., and Finkelstein, A. (1976). Reconstitution in Planar Lipid Bilayers of a Voltage-Dependent Anion-Selective Channel Obtained from Paramecium Mitochondria,J. Membr. Biol. 30 99–120.
Thinnes, F. P., Götz, H., Kayser, H., Benz, R., Schmidt, W. E., Kratzin, H. D., and Hilschmann, N. (1989). Reinigung Eines Porins aus Menschlichen B-Lymphozyten (Porin 31HL) und Sein Topochemischer Nachweis auf Dem Plasmalemm der Herkunftszelle,J. Biol. Chem. Hopper-Seyler 370 1253–1264.
Thomas, L., Kocsis, E., Colombini, M., Erbe, E., Trus, B. L., and Steven, A. C. (1991). Surface Topography and Molecular Stoichiometry of the Mitochondrial Channel, VDAC, in Crystalline Arrays.J. Struct. Biol. 106 161–171.
Weiss, M. S., Wacker, T., Weckesser, J., Welte, W., and Schulz, G. E. (1990). The Three-Dimensional Structure of Porin fromRhodobacter capsulatus at 3 Å Resolution,FEBS Lett. 267 268–272.
Zhang, D. W., and Colombini, M. (1990). Group III-Metal Hydroxides Indirectly Neutralize the Voltage Sensor of the Voltage-Dependent Mitochondrial Channel, VDAC, by Interacting with a Dynamic Binding Site,Biochim. Biophys. Acta 1025 127–134.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Peng, S., Blachly-Dyson, E., Colombini, M. et al. Determination of the number of polypeptide subunits in a functional VDAC channel fromSaccharomyces cerevisiae . J Bioenerg Biomembr 24, 27–31 (1992). https://doi.org/10.1007/BF00769527
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF00769527