Lab 10: PRELAB

PRELAB ASSIGNMENT

• What will you be doing in lab? Why?
• What are the concentrations of your wild type HCAII and mutant HCAII (in µM)?
• What range of substrate concentrations are you going to use to determine the K_M of your protein? If you use the enzyme database BRENDA, you can find the published K_M value for the wild type enzyme, which will help you plan your experiment (link opens in a new tab): http://www.brenda-enzymes.info/
• Sketch a Michaelis-Menton curve and label the V_max and K_M – what data will you need to collect during lab in order to determine the V_max and K_M of HCAII?
• Complete planning and calculations for the lab PRIOR to lab!
  • You must coordinate with your podmates
  • This video from a TA helps explain the planning process:

PRELAB QUIZ

As mentioned above, you can use the BRENDA database to estimate the expected K_M value of wt HCAII for PNPA (which is also called 4-NPA). Which of the following is approximately the expected K_M?

a) 2.4 nM

b) 2.9 mM

c) 15.4 mM

Please answer the following true/false question:

You can use either Sec^-1 or Min^-1 as the unit for k_cat.

Link to the prelab quiz in Canvas: Prelab 10 Quiz