Enzyme Kinetics

Question 1

What is the Michaelis-Menten equation?

Answer 1

Question 2

How can you calculate the max. velocity for a given enzymatic process given the concentration of an enzyme and the turnover number?

Answer 2

K_cat - “turnover number,” determine the rate an enzyme can convert substrate to product

Question 3

What is the mathematical and conceptual definitions of k_m?

Answer 3

K_m = [S] ∙ 1/2(V_max) and is used to determine the affinity of the enzyme to the substrate. Lower K_m values indicate increased affinity and vice versa, because:

Question 4

What is catalytic efficiency, and how is it calculated?

Answer 4

k_cat = k_cat/k_m

Which is the measure of an enzymes specificity, with low K,_m and large K,_cat values indicating an enzyme that is highly specific to a given substrate. Catalytic efficiency is concerned with the reaction rate at low substrate concentrations.

Question 5

What is the Hill Coefficient, and what information can be obtained from its numerical information?

Answer 5

“The Hill coefficient (n_H) is a central parameter in the study of ligand-protein interactions, which measures the degree of cooperativity between subunits that bind the ligand in multisubunit proteins.” -Hagai Abeliovich

When the Hill coefficient is larger than 1, this indicates positive cooperativity. This means that the binding of one effector/substrate will increase the binding affinity for another.

When the HIll coefficient is equal to 1, there is no cooperativity, and that specific effector/substrate has not impact on the binding affinity of subsequent effectors/substrates.

When the Hill coefficienty is less than 1, this indicates negative cooperativity. So that the binding of the effector/substrate lessens the binding affinity of subsequent effectors/substrates.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1366580/#:~:text=The%20Hill%20coefficient%20(nH,the%20ligand%20in%20multisubunit%20proteins.

Question 6

What type of inhibition does the following Lineweaver-Burk plot represent? What are the main characteristics?

Answer 6

Competitive Inhibition

Question 7

What type of inhibition does the following Lineweaver-Burk plot represent? What are the main characteristics?

Answer 7

Uncompetitive Inhibition

Question 8

What type of inhibition does the following Lineweaver-Burk plot represent? What are the main characteristics?

Answer 8

Noncompetitive Inhibition

Question 9

What is the conceptual defintion of noncompetitive inhibition?

Answer 9

“Non-competitive inhibition effectively reduces the amount of enzyme by the same fixed amount in a typical experiment at every substrate concentration used,” thus the V_max is reduced due to noncompetitive inhibition becuase V_max is dependent on the concentration of available enzyme.

https://chem.libretexts.org/Courses/University_of_Arkansas_Little_Rock/CHEM_4320_5320%3A_Biochemistry_1/05%3A_Michaelis-Menten_Enzyme_Kinetics/5.4%3A_Enzyme_Inhibition

Question 10

What is the conceptual defintion of uncompetitive inhibition?

Answer 10

An uncompetitive inhibitor binds allosterically to the ES complex, and locks the substrate into the active site to form the ESI complex. This type of inhibition effictely lowers both k_m and V_max.

Question 11

In regards to a Lineweaver-Burk plot, what type of inhibition will increase K_m while V_max is unchanged when compared to normal enzymatic activity?

Answer 11

Competitive inhibition

Question 12

In regards to a Lineweaver-Burk plot, what type of inhibition will decrease K_m while also decreasing V_max when compared to normal enzymatic activity?

Answer 12

Uncompetitive Inhibition

Question 13

In regards to a Lineweaver-Burk plot, what type of inhibition will decrease V_max while K_m is unchanged when compared to normal enzymatic activity?

Answer 13

Noncompetitive AKA Pure Noncompetitive

1/V_max is increased when a pure noncompetitive inhibitor is added, therefore; the maximum activity rate for the enzymatic process is decreased.

Question 14

In regards to a Lineweaver-Burk plot, what type of inhibition will decrease V_max while increasing K_m when compared to normal enzymatic activity?

Answer 14

Mixed Inhibition

Question 15

What is the slope of a Lineweaver-Burk plot equal to?

Answer 15

m = k_m/V_max

Question 16

What experimental conditions are necessary to achieve reliable data for Michaelis-Menten enzyme kinetics?

Answer 16

• Velocity of the reaction must be measured under steady state conditions
• Solution pH and temperature must remain constant at all substrate concentrations
• The concentration of the enzyme must be lower than that of the substrate (steady state kinetics)