Journal Club 3: Fields & Song, Nature, 1989
The authors state that “protein-protein interactions between two proteins have generally been studied using biochemical techniques such as cross-linking, co-immunoprecipitation and co-fractionation by chromatography.” Briefly describe one of these techniques.
What does the GAL4 protein normally do in Saccharomyces cerevisiae (Brewer’s yeast)?
What are the attributes of GAL4 that enable it to be used in the yeast two hybrid system?
What is GAL80 and why does the yeast two hybrid system require mutant strains of yeast that lack the expression of this protein?
What is the UASg sequence used for?
Why did the authors choose SNF1 and SNF4 as the first proteins to test in their system?
Below is a table of their results:
What do the numbers in parentheses represent?
What does the data in row 2 demonstrate?
What does the data in rows 8 and 9 demonstrate?
What row demonstrates the expected experimental results?
How was Beta-galactosidase activity measured?
The authors state that the system “requires that the interaction can occur within the yeast nucleus, that the GAL4-activating region is accessible to the transcription machinery, and that the GAL4(1-147)-protein X hybrid is itself not a potent activator.” Briefly describe why each of these requirements must be met for the system to properly function.