Enzymes Flashcards
1
Q
Enzyme
A
- Protein or RNA molecule that catalyzes biochemical reactions
- Bind specific substrates with high fidelity and specificity
- Enzymes catalyze reactions under mild conditions
- Most if not all metabolic pathways are regulated by regulating enzyme function
2
Q
How Do Enzymes Work
A
- Specific for substrates
- Increase reaction rates by increasing local concentrations of reactants and stabilizing the transition state
3
Q
Specificity
A
- Binding of substrate to active site initiates catalysis
- Binding site is complementary to substrate structure
- High specificity for substrates
- Some enzymes catalyze highly stereospecific reactions
- Shape must be right
4
Q
Models of Enzyme Substrate Interactions - Lock and key: DHFR
A
- Yellow DHF molecule is bound and then NADPH fits easily into the performed site
- Conformal changes in Dihydrofolate Reductase
5
Q
Models of Enzyme Substrate Interactions - Induced Fit
A
- Hexose Kinase
- Binding of hexose causes conformational change that allows active site to form
6
Q
How Catalysts Work
A
- Reduction of Activation Energy
- Delta Gº’ is NOT changed
- Catalyst is not consumed by reaction
- Increases local concentration of reactants
- Insure proper orientation of reactants
- Increase local concentrations of reactive groups
- Stabilize the transition state
7
Q
Cofactors
A
- Small molecules that provide additional reactive groups for catalysis
- NAD+/NADH
- Biotin
- Riboflavin
- Usually added to the diet as vitamins
8
Q
How To Lower Activation Energy
A
- Enzymes bind transition states best
- Enzyme active sites are complimentary to the transition state of the reaction
- Enzymes bind transition states better than substrates
- This lowers the ctivation barrier
- Enzyme uses binding energy of substrates to organize the reactants to a fairly rigid ES complex
- Entropy cost paid during binding, all interactions contribute to the binding energy
9
Q
Two things affected by enzyme
A
- Delta S of reaction
- Delta H of reaction
10
Q
Measurable Properties of an Enzyme
A
- Vmax, fastest a reaction can proceed in the presence of an enzyme
- Km, how the substrate binds to the enzyme
- Vmax/Km, how efficient the enzyme is
11
Q
Michaelis-Menten Plot
A
- Each point on curve represents one experimental point with the same E and different S
- Then measuring rate for that combination
- Measure Vmax and Km
12
Q
Michaelis-Menten Kinetics Assumptions
A
- Step 1 faster than Step 2
- Step 2 is rate limiting
- [ES] is at a steady state
- Initial rates are used so that [P] = 0, assume step 2 is irreversible
- [S] is much greater than [E]t, initially [S] does not change
- [E]t = [E] + [ES]. no alternate forms of the enzyme
13
Q
Km
A
- [S] at half maximal velocity
- Typical units of Km are mM or uM
- (k-1 + k2) / k1
- If Km is large, substrate does not bind well to the enzyme, k2 is large
- If Km is small, substrate binds well to enzyme, k2 is small
14
Q
Vmax
A
- Maximal velocity for a given [E]t
- Typical units are umoles of product per unit time
- Vmax = k2[E]t
15
Q
k2/Km
A
- Measure of catalytic efficiency
- smallest km not necessarily best enzyme
16
Q
Determining Vmax and Km
A
- Best method is to fit the V vs [S] data to the Michaelis-Menten equation directly
- Linearize the equation
17
Q
Lineweaver-Burk Plot
A
- Line
- 1/[S] on the x-axis
- 1/[v] on the y-axis
- Slope of Km/Vmax
- y-intercept of 1/Vmax
- Best way to study inhibitors
18
Q
Enzyme Inhibitors Info
A
- Billion dollar industry
- Most drugs are enzyme inhibitors
- HIV treatments include protease inhibitors and reverse transcriptase inhibitors
19
Q
How Enzyme Inhibition works
A
Slow down the activity of an enzyme
20
Q
Competitive Inhibition
A
- Binds to the active site in place of the substrate and can only bind in absence of substrate
- At high enough [S] the substrate out competes the inhibitor
- Increases Km, doesn’t change Vmax
21
Q
Uncompetitive Inhibition
A
- Binds to another site and can bind only in the presence of substrate
- Inhibitor can only interact with S at the active site
- Km is decreased and Vmax is decreased
22
Q
Mixed Inhibition
A
- Binds to another site and can bind in the presence or absence of substrate
- Vmax decreases and Km can increase or decreases
23
Q
Irreversible Inhibitors
A
- Form covalent adducts with the enzyme
- DIFP and TCPK are irreversible inhibitors of the serine proteases
