PharmSci IV Exam 3 Flashcards
(90 cards)
1
Q
In the Michaelis-Menten Enzyme kinetics model, describe the reaction velocity at different concentrations of substrate. (2 components)
A
- At low [S], velocity is proportional to [S].
2. At high [S], reaction is independent of [S] and plateaus.
2
Q
Why is it useful to define the Michaelis-Menten constant, Km? (2 components)
A
- [ES] is a transient species, that cannot be measured, dependent on reaction rates of association and dissociation. Km contains these reaction rates
- With steady-state assumption, [ES] = [E][S]/Km.
3
Q
What is the simplified Michaelis-Menten equation?
A
V = Vmax * ([S]/[S] + Km])
4
Q
How does the simplified Michael-Menten equation fit the observations of enzyme reaction velocity at different concentrations of substrate? (2 components)
A
- When Km»_space; [S], equation simplifies to V = (Vmax/Km)[S], e.g. it is proportional to [S].
- When [S]»_space; Km, equation simplifies to V = Vmax, e.g. reaction plateaus.
5
Q
State the Lineweaver-Burke Equation for Enzyme kinetics.
A
1/V = (1/Vmax) + (Km/Vmax)*(1/[S])
6
Q
State the values of the slope, y-intercept and x-intercept of the Lineweaver-Burke Equation. (3 components)
A
- Slope = (Km/Vmax)
- Y-intercept = (1/Vmax)
- X-intercept = -1/Km.
7
Q
How is Lineweaver-Burke plot used experimentally? (3 steps)
A
- Measure V for different [S]
- Plot 1/V vs. 1/[S]
- Use slope, y-intercept and x-intercept to calculate Km and Vmax in order to characterize the enzyme.
8
Q
What is the physical significance of Km? (2 components)
A
- Using Lineweaver-Burke equation, when [S] = Km, V = Vmax/2. Thus Km is the [S] at which velocity is half of maximum.
- Therefore, an enzyme with a small Km reaches maximum catalytic activity at a low [S]. This makes it more efficient. Thus Km is a means to compare the [S] required for efficient catalysis… often this means it is a measure of affinity for substrate. (Remember: Small Km = High Affinity)
9
Q
Define Kcat
A
Kcat = Vmax / [total enzyme]
10
Q
What is the physical significance of Kcat?
A
It is the number of substrate molecules that undergo catalytic production per unit time (seconds). It is the “turnover number.”
11
Q
What is the purpose of Kcat/Km?
A
- A measure of catalytic efficiency.
2. This number is high when there is a large turnover (large Kcat) or high affinity (small Km).
12
Q
Define Enzyme Unit (U)
A
An enzyme unit is that which catalyzes that conversion of one mMol of substrate per minute.
13
Q
Describe and define specific enzymatic activity (2 components)
A
- Specific activity relates the Enzyme Unit (a measure of activity) to the total Amount of Enzyme.
- Define as Enzyme Unit / mg protein.
14
Q
What are seven (7) factors affecting enzyme activity in vivo?
A
- Genetic
- Age
- Sex
- Nutrition and Disease
- Enzyme activation/inhibition
- Enzyme induction
- Species differences
15
Q
Define an isoenzyme
A
Different amino acid sequence (different genes), same chemical reaction (substrate).
16
Q
Define an isoform
A
Same gene, but different ultimate protein structure (through RNA splicing mechanisms)
17
Q
Describe enterohepatic cycling? (4 components)
A
A circuit from:
- Liver
- Bile
- Small Intestine… reabsorption
- Liver
18
Q
What is the net effect of enterohepatic cycling?
A
- Drugs and other substances (e.g. bile) may stay within the enterohepatic cycle for a long time
- Unexpected changes in plasma concentration may be seen as a result of cycling
19
Q
What are three (3) classes of enzymes involved with biotransformation?
A
- Xenobiotic-transforming enzymes
- Endobiotic transforming enzymes
- Microflora enzymes
20
Q
What tissues express enzymatic proteins?
A
Virtually all tissues.
21
Q
Where does most xenobiotic metabolism occur?
A
Endoplasmic reticulum of liver.
22
Q
What are four (4) common organelles that contain enzymatic proteins?
A
- Microsomes (endoplasmic reticulum)
- Cytosol
- Plasma membrane
- Inner mitochondrial membrane
23
Q
What are five (5) major factors affecting drug metabolism?
A
- Genetics
- Physiologic factors (e.g. age, gender, microflora)
- Pharmacodynamic (e.g. dose, frequency, route)
- Environmental factors (e.g. pesticides)
- Transporter proteins
24
Q
What are five (5) models for studying metabolism/enzymes?
A
- Whole animal
- Organ/Cell
- Cell fractions (subcellular)
- Purified enzymes
- X-ray crystallography
25
What is first-pass metabolism?
Metabolism that occurs from by small intestine and liver before entering systemic circulation.
26
What is the mechanism of monooxygenase?
1. Two atoms of oxygen are reduced to -OH and -H2O (from O2)
| 2. -OH is added to the substrate.
27
What are the enzymes and cofactors for hydroxylation?
CYP, NADPH, H+, O2
28
What are the products for aromatic hydroxylation? (2 components)
1. arene oxide
| 2. phenol
29
What are the products for aliphatic hydroxylation? (2 components)
1. If alkane, then alcohol.
| 2. If alkene, then epoxide.
30
What are the enzymes and cofactors for dealkylation?
CYP, NADPH, H+, O2
31
What are the products of N-dealkylation?
An amine + (aldehyde or ketone.)
32
What is the intermediate for N-dealkylation?
Hemiaminal... put an -OH on the C of N-C.
33
What are the products of O-Dealkylation?
(An alcohol or phenol) + (aldehyde or ketone.)
34
What is the intermediate for O-Dealkylation?
Hemiacetal... put an -OH on the C of O-C.
35
What are the enzymes and cofactors for N-oxidation?
(CYP or FMO), NADPH, H+, O2
1. CYP prefers amides and primary amines
2. FMO prefers secondary and tertiary amines
36
What is the product of N-oxidation?
NH converted to N-OH
37
What are the enzymes and cofactors for S-oxidation?
(Primarily FMO, or CYP), NADPH, H+, O2
38
What are the products of S-oxidation?
Add =O to sulfur; can also have O=S=O
39
What are the products of Beta-oxidation of carboxylic acids?
Cleave at the Beta carbon and form two carboxylic acids.
40
What are the enzymes and cofactors for purine and pyrimidine oxidation?
Xanthine oxidase
41
What is the product of purine and pyrimidine oxidation?
Add -OH between N of each ring; one on each ring.
42
What are the enzymes and cofactors for oxidative desulfuration?
CYP, NADPH, H+, O2
43
What are the products for oxidative desulfuration?
Replace =S with =O
44
What are the enzymes and cofactors for dehalogenation?
CYP, NADPH, H+, O2
45
What are the products of dehalogenation?
Replace HX with =O; e.g. CHCl3 becomes COCl2 (phosgene)
46
What are the enzymes and cofactors for deamination?
Monoamine oxidase, O2
47
What are the products of deamination?
A primary amine becomes an (aldehyde or ketone) + NH3 + H2O2
48
What are the enzymes and cofactors for alcohol oxidation?
Alcohol dehydrogenase (ADH), NAD+
49
What are the products of alcohol oxidation?
Alcohol becomes (aldehyde or ketone)
50
What are the enzymes and cofactors for aldehyde oxidation?
Aldehyde dehydrogenase, NAD+
51
What are the products of aldehyde oxidation?
Aldehyde becomes carboxylic acid.
52
What are the enzymes and cofactors for dihydrodiol oxidation?
Dihydrodiol dehydrogenase, NADP+
53
What are the products for dihydrodiol oxidation?
A quinone
54
What are the enzymes and cofactors for aldehyde/ketone reduction?
carbonyl reductase, NADPH
55
What are the products of aldehyde/ketone reduction?
(Aldehyde or ketone) becomes an alcohol
56
What are the enzymes and cofactors for azo reduction?
gut CYP or microflora enzymes
57
What are the products of azo reduction?
R-N=N-R' becomes R-NH2 and R'-NH2
58
What are the enzymes and cofactors for nitro reduction?
gut CYP or microflora enzymes
59
What are the products of nitro reduction?
R-NO2 becomes R-NH2
60
What are the enzymes and cofactors for quinone reduction?
Quinone reductase
61
Why is quinone dangerous in the body?
1. One-electron reductino of quinone yields semiquinone radical.
2. Semiquinone radical reacts with O2 to form superoxide radical which damages DNA.
62
What are the enzymes and cofactors for epoxide hydrolysis?
Epoxide hydrolase, H2O
63
What are the products of epoxide hydrolysis?
An epoxide becomes a diol.
64
What are the enzymes and cofactors for ester hydrolysis?
Esterases, H2O
65
What are the products of ester hydrolysis?
An ester becomes an acid + alcohol.
66
What are the products of amide hydrolysis?
An amide becomes an acid + amine.
67
What is transesterification?
Esterase enzymes can catalyze the transfer of alkyl groups between an ester and an alcohol.
68
What are the enzymes and cofactors involved with glucoronidation?
UGT, UDPGA
69
What are the acceptors for glucoronidation? (4 components)
Nucleophiles
1. R-OH
2. R-NH2
3. Acids
4. R-SH
70
Describe the activity of glucoronidation in terms of affinity and capacity
Low affinity, high capacity.
71
What are the enzymes and cofactors involved with sulfation?
Sulfotransferase (SULT), PAPS
72
What are the acceptors of sulfation?
R-OH
73
Describe the activity of sulfation in terms of affinity and capacity
High affinity, low capacity
74
What are the enzymes and cofactors involved with acetylation?
N-Acetyl Transferase (NAT), acetyl CoA
75
What are the acceptors of acetylation?
Primary amines, -NH2 becomes -NH-CCH3=O
76
What are the enzymes and cofactors involved with glycine conjugation?
CoA, Glycine
77
What are the acceptors for glycine conjugation?
Carboxylate (carboxylic acid) accepts glycine and forms amide.
78
What are the enzymes and cofactors involved with glutathione conjugation?
Glutathione-S-Transferase (GST), Glutathione (GSH)
79
What is the amino acid structure of Glutathione?
Gamma-Glu-Cys-Gly
80
What are the acceptors for glutathione conjugation? (4 components)
Electrophiles
1. Halides
2. Epoxides
3. Unsatured carbonyls
4. Quinones
81
What enzyme is involved with the conjugation of glutathione to peroxide?
Glutathione peroxidase
82
What occurs after glutathione is conjugated to a substrate? (2 components)
1. Undergoes processing to form a mercapturic acid
| 2. Excreted
83
What enzymes and cofactors are involved with methylation?
Methyl transferase, SAM
84
What chemical structures are dangerous in the body? (4 components)
1. Quinones
2. Aldehydes/carbonyls/ketones
3. Epoxides
4. Unsaturated carbonyls
85
What is hyperplasia?
Uncontrolled cell growth
86
What is anaplasia?
Lack of differentiation [resembles embryonic cells]
87
What is metastasis?
Movement of tumor from primary to secondary site [characterizes malignant tumor]
88
What are three ways carcinogens can cause cancer?
1. Damage to oncogenes, resulting in uncontrolled cell proliferation
2. Damage to tumor suppressor genes
3. Loss of function of DNA repair genes/proteins
89
What oxidation reactions involve FMO enzyme?
N-oxidation of substitute amines, S-oxidation
90
Describe the difference between FMO and CYP intermediates
1. CYP has a heme with activated oxygen intermediate
| 2. FMO has a flavin hydroperoxide intermediate
