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The F1 ATP synthetase beta-subunit: a major yeast novobiocin binding protein
J R Jenkins1, M J Pocklington, E Orr
1Department of Genetics, University of Leicester, England.
Abstract:
Novobiocin affects DNA metabolism in both prokaryotes and eukaryotes, resulting in cell death. In prokaryotes, the drug is a specific inhibitor of DNA gyrase, a type II topoisomerase that can be purified on a novobiocin-Sepharose column. The yeast type II topoisomerase is neither the biochemical, nor the genetic target of the antibiotic. We have purified the major yeast novobiocin binding proteins and identified one of them as the beta-subunit of the yeast mitochondrial F1 ATP synthetase, a protein highly conserved throughout evolution. The inactivation of this protein might explain the toxic effects of novobiocin on higher eukaryotic cells.
Insights
Novobiocin kills cells by impacting DNA metabolism. In yeast, it doesn't target DNA gyrase but binds to mitochondrial F1 ATP synthase's beta-subunit, explaining its toxicity.
Area of Science:
- Molecular Biology
- Biochemistry
- Cell Biology
Background:
- Novobiocin is known to affect DNA metabolism and cause cell death in prokaryotes and eukaryotes.
- In prokaryotes, novobiocin specifically inhibits DNA gyrase, a type II topoisomerase.
- The mechanism of novobiocin's toxicity in eukaryotes, particularly yeast, was not well understood.
Purpose of the Study:
- To investigate the biochemical target of novobiocin in yeast.
- To identify the proteins responsible for novobiocin binding in eukaryotic cells.
- To elucidate the mechanism behind novobiocin's toxic effects on higher eukaryotic cells.
Main Methods:
- Purification of major novobiocin binding proteins from yeast.
- Biochemical assays to determine protein-drug interactions.
- Identification of purified proteins using established molecular biology techniques.
Main Results:
- Yeast type II topoisomerase was confirmed not to be the biochemical or genetic target of novobiocin.
- The major novobiocin binding proteins in yeast were successfully purified.
- One key novobiocin binding protein was identified as the beta-subunit of yeast mitochondrial F1 ATP synthetase.
Conclusions:
- Novobiocin's mechanism of action in yeast differs from its prokaryotic target, DNA gyrase.
- The beta-subunit of mitochondrial F1 ATP synthetase is a primary novobiocin binding protein in yeast.
- Inactivation of this highly conserved mitochondrial protein likely underlies novobiocin's toxicity in higher eukaryotes.