2-7 Enzymes as Biological Catalysts

Question 1

What are the general characteristics of enzymes?

Answer 1

• Carry out almost all chemistry required by living systems
• Permit a wide range of chemical reactions in a narrow set of conditions
• Allow rapid, efficient adjustments to environmental conditions
• Enhance reaction rate tremendously
• Require extreme specificity
• Capable of being regulated

Question 2

What are the common features of active sites on enzymes?

Answer 2

• Occupy a small part of total volume of most enzymes
• 3-D structure
• Bind substrates through multiple weak, non-covalent interactions (electrostatic, H bonds, van der Waals, hydrophobic effect)
• Water is excluded unless it is a reagent (active sites are in clefts of proteins)
• Highly specific binding of substrate
• Can include non-protein prosthetic groups and cofactors

Question 3

What is the model for an enzymatic reaction?

Answer 3

(Formation of the enzyme-substrate complex involves induced fit)

Question 4

What is the Michaelis-Menten equation?

Answer 4

Question 5

What does K_m mean in the context of the Michaelis-Menten reaction?

Answer 5

• K_m = [S] at which v = 1/2 V_max
• K_m = ((k₂ + k₃) / k₁)
• The catalytic rate of an enzyme is most sensitive to [S] when [S] << K_m
• Knowing K_m allows you to calculate what fraction of enzyme molecules are bound to substrate at any [S]:
• *f_ES = (v / V_max) = ([S] / ([S] + K_m))**
• When k₂ >> k₃ (i.e., the substrate binds and dissociates many times before it goes on to form product), K_m ≈ K_d (the dissociation constant for the ES complex)
• k₂ >> k₃ is relatively common; under these conditions, K_m gives you an idea of the affinity of enzyme for substrate: high K_m = low affinity, low K_m = high affinity

Question 6

What does V_max mean in the context of the Michaelis-Menten equation?

Answer 6

V_max = maximal velocity achievable for a specific concentration of enzyme

Although increasing [S] does not give further increases in initial velocity at V_max, increasing [E] will increase the V_max. (At V_max, enzyme binding sites are saturated with substrate, so increasing [E] can relieve that saturation.)

Question 7

What does k_cat mean in the context of the Michaelis-Menten equation?

Answer 7

AKA k₃, or the “turnover number” because it describes the number of substrate molecules that can be “turned over” (converted to product) by a single enzyme molecule in a given period of time

k_cat = V_max / [E]_t
(where [E]_t = total enzyme concentration)

Question 8

How can the Michaelis-Menten equation constants be used to provide a comparison of enzyme efficiencies?

Answer 8

k_cat/K_m

A higher k_cat/K_m implies higher efficiency, because k_cat determines how quickly the ES compex is used and K_m describes how much ES complex is available.

Question 9

What is a Lineweaver-Burk plot?

Answer 9

A linearized form of the Michaelis-Menten equation. Can be useful with enzyme inhibitors.

Question 10

What are the different types of enzyme inhibition?

Answer 10

• Irreversible: bind very tightly to an enzyme and inactivate an essential functional group
• Reversible: bind reversibly to an enzyme and temporarily inactivate it while bound

1. Competitive: compete with substrate for binding to the active site; can be overcome by increasing [S]; K_m appears to increase, but V_max does not change (on L-B plot, same y-intercept but different x-intercepts)
2. Non-competitive: bind to a site on the enzyme OTHER than the active site; CANNOT be overcome by increasing [S]; V_max appears to increase, but K_m does not change (on L-B plot, same x-intercept, but different y-intercepts)
3. Uncompetitive

Question 11

What does K_i mean with regard to enzyme inhibition?

Answer 11

K_i is the dissociation constant for the enzyme-inhibitor complex. It describes the strength with which an inhibitor binds to an enzyme.

Higher K_i implies weaker inhibition.

Question 12

How can Lineweaver-Burk plots be used to calculate K_i?

Answer 12

• (slope in the presence of inhibitor / slope in the absence of inhibitor) = α
• α = 1 + ( [I] / K_i )
• Solve for K_i