Diiron centre mutations in Ciona intestinalis alternative oxidase abolish enzymatic activity and prevent rescue of cytochrome oxidase deficiency in flies

Sci Rep. 2015 Dec 17;5:18295. doi: 10.1038/srep18295.

Abstract

The mitochondrial alternative oxidase, AOX, carries out the non proton-motive re-oxidation of ubiquinol by oxygen in lower eukaryotes, plants and some animals. Here we created a modified version of AOX from Ciona instestinalis, carrying mutations at conserved residues predicted to be required for chelation of the diiron prosthetic group. The modified protein was stably expressed in mammalian cells or flies, but lacked enzymatic activity and was unable to rescue the phenotypes of flies knocked down for a subunit of cytochrome oxidase. The mutated AOX transgene is thus a potentially useful tool in studies of the physiological effects of AOX expression.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Animals, Genetically Modified
  • Cell Line
  • Ciona intestinalis / enzymology*
  • Ciona intestinalis / genetics
  • Drosophila Proteins / deficiency
  • Drosophila Proteins / genetics
  • Drosophila melanogaster / cytology
  • Drosophila melanogaster / enzymology*
  • Drosophila melanogaster / genetics
  • Electron Transport Complex IV / genetics
  • Electron Transport Complex IV / metabolism*
  • Female
  • Gene Knockdown Techniques
  • HEK293 Cells
  • Humans
  • Iron / chemistry
  • Iron / metabolism*
  • Male
  • Mitochondrial Proteins / chemistry
  • Mitochondrial Proteins / genetics
  • Mitochondrial Proteins / metabolism*
  • Models, Molecular
  • Molecular Sequence Data
  • Mutation*
  • Oxidoreductases / chemistry
  • Oxidoreductases / genetics
  • Oxidoreductases / metabolism*
  • Oxygen Consumption
  • Plant Proteins / chemistry
  • Plant Proteins / genetics
  • Plant Proteins / metabolism*
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid

Substances

  • Drosophila Proteins
  • Mitochondrial Proteins
  • Plant Proteins
  • Iron
  • Oxidoreductases
  • alternative oxidase
  • Electron Transport Complex IV