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Botany JoVE (Open Access) Citable · DOI

Tuning Degradation to Achieve Specific and Efficient Protein Depletion

DOI: 10.3791/59874-v
What you'll learn
  • ✓ Apply the β-est AID system to deplete target proteins in S. cerevisiae
  • ✓ Tune degradation rates for specific and efficient protein depletion
  • ✓ Analyze and quantify protein depletion kinetics using experimental readouts
Protocol

Here, we present a protocol to effectively and specifically deplete a protein of interest in the yeast Saccharomyces cerevisiae using the β-est AID system.

Difficulty
advanced
Total time
~4–8 hours per experiment (induction to analysis)
Model organism
Saccharomyces cerevisiae
Biosafety
BSL-1

Steps

1
Execute general procedure for protein depletion

Apply the β-est AID system to induce specific degradation of the target protein in yeast cells. Follow the protocol to initiate and monitor protein depletion over time.

▶ 00:36
2
Analyze tuned depletion rate kinetics

Measure and quantify the degradation kinetics of the depleted protein to confirm specificity and efficiency. Compare depletion profiles across different tuning conditions.

▶ 03:31
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