Purification and characterization of the voltage-dependent anion channel from the outer mitochondrial membrane of yeast.
The outer mitochondrial membranes of all organisms so far examined contain a protein which forms voltage-dependent anion selective channels (VDAC) when incorporated into planar phospholipid membranes. Previous reports have suggested that the yeast (Saccharomyces cerevisiae) outer mitochondrial membrane component responsible for channel formation is a protein of 29,000 daltons which is also the major component of this membrane. In this report, we describe the purification of this 29,000-dalton protein to virtual homogeneity from yeast outer mitochondrial membranes. The purified protein readily incorporates into planar phospholipid membranes to produce ionic channels. Electrophysiological characterization of these channels has demonstrated they have a size, selectivity and voltage dependence similar to VDAC from other organisms. Biochemically, the purified protein has been characterized by determining its amino acid composition and isoelectric point (pI). In addition, we have shown that the purified protein, when reconstituted into liposomes, can bind hexokinase in a glucose-6-phosphate dependent manner, as has been shown for VDAC purified from other sources. Since physiological characterization suggests that the functional parameters of this protein have been conserved, antibodies specific to yeast VDAC have been used to assess antigenic conservation among mitochondrial proteins from a wide number of species. These experiments have shown that yeast VDAC antibodies will recognize single mitochondrial proteins from Drosophila, Dictyostelium and Neurospora of the appropriate molecular weight to be VDAC from these organisms. No reaction was seen to any mitochondrial protein from rat liver, rainbow trout, Paramecium, or mung bean. In addition, yeast VDAC antibodies will recognize a 50-kDa mol wt protein present in tobacco chloroplasts.(ABSTRACT TRUNCATED AT 250 WORDS)
D000596:Amino Acids / Q000032:analysis; D006593:Hexokinase / Q000378:metabolism; D007425:Intracellular Membranes / Q000502:physiology*; D007473:Ion Channels / Q000502:physiology*; D008051:Lipid Bilayers; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D013367:Submitochondrial Particles / Q000502:physiology*
D000596:Amino Acids; D007473:Ion Channels; D008051:Lipid Bilayers; D008565:Membrane Proteins; D006593:Hexokinase