ASSIGNMENT
Use chromatography to purify a protein of interest.
Use a colorimetric assays to examine protein activity and folding.
Determine the inhibition of several antibiotics with the protein.
Determine how various reagents alter the stability/activity of the protein.
Examine protein purity using gel electrophoresis.
Generate a protein concentration standard curve and calculate the concentration of your protein solutions.
Use PyMol to visualize β-lactamase proteins.
What are the different methods used to purify proteins from crude extract materials? Why are so many different types of methods required?
What is happening in each step of our purification protocol?
How does gel electrophoresis work? What characteristics of the protein determine how far it will migrate in a gel? We will run denaturing gels, what does this mean and why is it important?
How do various reagents, such as urea, hydrogen peroxide and change in pH, affect protein activity and stability?
What is a protein’s “specific activity”?
What is the basis for the nitrocefin-based activity assay? How does it work?
How does Bis-ANS enable visualization of protein folding?
