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Cycloheximide Chase Analysis of Protein Degradation in Saccharomyces cerevisiae
Published on: April 18, 2016
Cell cycle-dependent protein secretion by Saccharomyces cerevisiae.
1Department of Chemical Engineering and Materials Science, 151 Amundson Hall, 421 Washington Ave. S.E., Minneapolis, MN 55455-0132, USA.
Biotechnology and Bioengineering
|October 23, 2001
Summary
Synchronizing yeast cells (Saccharomyces cerevisiae) revealed cell cycle-dependent protein secretion. Young daughter cells secrete less protein than older mother cells, impacting secretion rate studies.
Area of Science:
- Biochemistry
- Cell Biology
- Microbiology
Background:
- Understanding protein secretion in yeast is crucial for biotechnology.
- Cell cycle position can influence cellular processes, including secretion.
- Accurate synchronization methods are essential for reliable biological data.
Purpose of the Study:
- To investigate the impact of cell cycle position on heterologous protein secretion rates in Saccharomyces cerevisiae.
- To compare different cell synchronization methods and their effects on physiological state and data quality.
- To determine if young daughter cells exhibit different secretion behaviors compared to older mother cells.
Main Methods:
- Synchronization of Saccharomyces cerevisiae cell populations using centrifugal elutriation.
- Comparison of cell synchrony induced by elutriation versus cell cycle-arresting drugs.
- Measurement of heterologous protein secretion rates at different cell cycle stages.
- Analysis of physiological perturbation caused by synchronization techniques.
Main Results:
- Cell cycle-arresting drugs caused significant physiological perturbation, obscuring secretion data.
- Centrifugal elutriation yielded synchronized first-generation daughter cells with minimal physiological impact.
- Synchronized cells showed significant protein secretion only at cell division, indicating cell cycle dependence.
- Maximum secretion rate in synchronized cultures (7-14 molecules/cell/second) was lower than in asynchronous cultures (29-51 molecules/cell/second).
Conclusions:
- Protein secretion in Saccharomyces cerevisiae is strongly dependent on cell cycle position.
- Young daughter cells exhibit distinct and lower protein secretion rates compared to older mother cells.
- Centrifugal elutriation is a preferred method for synchronizing yeast cells for secretion studies due to minimal physiological perturbation.
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