Podcast
Questions and Answers
One advantage of a Y2H screen is that it is a relatively fast and easy way to screen for ______ interactions
One advantage of a Y2H screen is that it is a relatively fast and easy way to screen for ______ interactions
protein-protein
Y2H requires little hands-on time and technical skill, making it ______ to use
Y2H requires little hands-on time and technical skill, making it ______ to use
easy
A limitation of Y2H screens is the high rates of false positives and false ______
A limitation of Y2H screens is the high rates of false positives and false ______
negatives
For the reporter gene to be activated in Y2H, the interaction must occur in the ______
For the reporter gene to be activated in Y2H, the interaction must occur in the ______
Signup and view all the answers
Protein folding, expression, and post-translational modifications may differ between yeast and humans, affecting their physical ______
Protein folding, expression, and post-translational modifications may differ between yeast and humans, affecting their physical ______
Signup and view all the answers
Tagged proteins in Y2H may not fold correctly, leading to abnormal binding of their ______
Tagged proteins in Y2H may not fold correctly, leading to abnormal binding of their ______
Signup and view all the answers
_______ roles of PPIs in cellular functions make them potential therapeutic targets.
_______ roles of PPIs in cellular functions make them potential therapeutic targets.
Signup and view all the answers
Targeted modulation of PPIs with small molecules is a promising approach with more than 50 PPIs successfully targeted with _______ molecules.
Targeted modulation of PPIs with small molecules is a promising approach with more than 50 PPIs successfully targeted with _______ molecules.
Signup and view all the answers
Most of the targeted PPIs are _______ of PPIs in drug development.
Most of the targeted PPIs are _______ of PPIs in drug development.
Signup and view all the answers
_______ of PPIs is also a strategy for targeted PPI modulation.
_______ of PPIs is also a strategy for targeted PPI modulation.
Signup and view all the answers
Yeast two-hybrid (Y2H) assay is an established technique to detect protein-protein interactions, and it is often used alongside one or two other methods to examine the _______ of interactions in cells.
Yeast two-hybrid (Y2H) assay is an established technique to detect protein-protein interactions, and it is often used alongside one or two other methods to examine the _______ of interactions in cells.
Signup and view all the answers
The yeast two-hybrid (Y2H) experiment detects physical interactions of proteins through the downstream activation of a _______ gene.
The yeast two-hybrid (Y2H) experiment detects physical interactions of proteins through the downstream activation of a _______ gene.
Signup and view all the answers
The Y2H assay relies on the expression of a reporter gene activated by the binding of a particular ______
The Y2H assay relies on the expression of a reporter gene activated by the binding of a particular ______
Signup and view all the answers
The transcription factor is comprised of a DNA-binding domain (BD) and an activation domain (AD). The protein of interest fused with the BD is known as the ______
The transcription factor is comprised of a DNA-binding domain (BD) and an activation domain (AD). The protein of interest fused with the BD is known as the ______
Signup and view all the answers
In order to activate the reporter gene expression, a transcriptional unit must be present at the gene locus, which is only possible if Bait and Prey ______
In order to activate the reporter gene expression, a transcriptional unit must be present at the gene locus, which is only possible if Bait and Prey ______
Signup and view all the answers
Expression of a reporter gene requires the binding of a transcription factor, which normally consists of two functionally and structurally independent domains: DNA-binding (DB) and activation (AD) ______
Expression of a reporter gene requires the binding of a transcription factor, which normally consists of two functionally and structurally independent domains: DNA-binding (DB) and activation (AD) ______
Signup and view all the answers
In the absence of Bait-Prey interaction, the AD domain is unable to localize to the reporter gene to drive gene ______
In the absence of Bait-Prey interaction, the AD domain is unable to localize to the reporter gene to drive gene ______
Signup and view all the answers
When Bait and Prey interact, the DB domain binds to the DNA localizing the AD domain upstream of the reporter gene, leading to the expression of reporter ______
When Bait and Prey interact, the DB domain binds to the DNA localizing the AD domain upstream of the reporter gene, leading to the expression of reporter ______
Signup and view all the answers
Study Notes
Advantages and Limitations of Y2H Screens
- A Y2H screen is a fast and easy way to screen for protein-protein interactions, requiring little hands-on time and technical skill.
- A limitation of Y2H screens is the high rates of false positives and false negatives.
Requirements for Y2H Assay
- The interaction must occur in the nucleus for the reporter gene to be activated in Y2H.
- Protein folding, expression, and post-translational modifications may differ between yeast and humans, affecting their physical interactions.
- Tagged proteins in Y2H may not fold correctly, leading to abnormal binding of their partners.
Y2H Assay Components
- The Y2H assay detects physical interactions of proteins through the downstream activation of a reporter gene.
- The assay relies on the expression of a reporter gene activated by the binding of a particular transcription factor.
- The transcription factor is comprised of a DNA-binding domain (BD) and an activation domain (AD).
- The protein of interest fused with the BD is known as the bait.
Mechanism of Y2H Assay
- Expression of a reporter gene requires the binding of a transcription factor, which normally consists of two functionally and structurally independent domains: DNA-binding (DB) and activation (AD) domains.
- In the absence of Bait-Prey interaction, the AD domain is unable to localize to the reporter gene to drive gene expression.
- When Bait and Prey interact, the DB domain binds to the DNA localizing the AD domain upstream of the reporter gene, leading to the expression of reporter genes.
Protein-Protein Interactions
- Modulation of protein-protein interactions (PPIs) with small molecules is a promising approach with more than 50 PPIs successfully targeted with small molecules.
- Most of the targeted PPIs are involved in drug development.
- Inhibition of PPIs is also a strategy for targeted PPI modulation.
Studying That Suits You
Use AI to generate personalized quizzes and flashcards to suit your learning preferences.
Description
Learn about the advantages and limitations of utilizing the Yeast Two-Hybrid (Y2H) screen for screening protein-protein interactions. Understand the speed, ease, scalability, as well as the potential drawbacks of using this technique.
