Enzymes Flashcards
(104 cards)
1
Q
Define “enzymes”
A
Protein catalysts of biologic origin which enhance the rates of biochemical reactions at a rate as to be compatible with life
2
Q
What are the 5 general properties of enzymes?
A
- Not altered or consumed during the reaction
- Only small amounts of enzyme are required
- Enzymes accelerate the speed at which a chemical reaction reaches equilibrium, but does not alter the equilibrium constant
- Each enzyme is highly specific for a given reaction; they act on only one substrate
- Enzymes act by lowering activation energy
3
Q
Occurs when the 3D structure begins to uncoil
A
Denaturation
4
Q
High or low (blank) causes denaturation in enzymes
A
Temperature and pH
5
Q
What is a cofactor?
A
Non-protein compounds required by some enzymes to make them active
6
Q
What is an inorganic cofactor?
A
Activator
7
Q
What is an organic cofactor?
A
Coenzyme
8
Q
Complete cofactor: enzyme complex
A
Holoenzyme
9
Q
What is a bound cofactor called?
A
Prosthetic group
10
Q
The protein portion of the enzyme is the ____
A
Apoenzyme
11
Q
A unique sequence and orientation of amino acids to form a pocket or groove that provides for the enzyme’s specificity for only a unique substrate
A
Active site
12
Q
A region other than the active site where a separate compound reacts, altering the shape of the active site, altering its fit w/ the substrate; is used to regulate enzyme activity
A
Allosteric site
13
Q
Different physical forms of an enzyme that all catalyze the same reaction
A
Isoenzymes
14
Q
The quantity of an enzyme that will catalyze the reaction of one micromole of substrate per minute under defined conditions
A
International unit
15
Q
Amount of enzyme catalyzed w/ a reaction rate of one mole per second
A
Katal
16
Q
The rate of enzymatic activity increases as the concentration of substrate increased until the maximum velocity of the reaction is achieved. This max is called the _____?
A
Vmax
17
Q
Half of the Vmax
A
Km
18
Q
List examples of activators (inorganic cofactors) and coenzymes (organic cofactors)
A
Inorganic: Zn2+, Fe2+, Cu2+, Mg2+, Mn2+
Organic: NAD+, NADH, NADP+, NADPH, pyrixodal-5-phosphate
19
Q
Why is enzyme activity, rather than ezyme concentration, measured in the lab?
A
The amount of enzyme is so small, that it’s difficult to devise assays sensitive enough. Instead, the rate of product formed or amount of substrate consumed during reaction is measured. The more enzyme present, the faster the reaction proceeds.
20
Q
Initial period when enzyme and substrate are first mixed, but no product is formed yet, so there is no detectable change in absorbance
A
Phases of an enzymatic reaction:
lag phase
21
Q
Rate of product formed is linear w/ time, so reaction follows Beer’s Law
A
Phases of an enzymatic reaction:
log (linear) phase
22
Q
No change in absorbance b/c there is no substrate left to be converted to product. Rate of reaction is dependent on substrate concentration not enzyme concentration.
A
Phases of enzymatic reaction:
Substrate depletion phase
23
Q
Why should enzyme measurements be made during the log phase?
A
It corresponds to zero order kinetics! It follows Beers Law so it is an accurate measurement
24
Q
For a typical enzyme measurement curve, how are the axis labeled?
A
X: time (seconds)
Y: change in absorbance
25
Enzyme Curve Zero order kinetics
- Define
- Where on graph
- The rate of the reaction is independent of reactant (substrate)
- Corresponds to Log (linear) phase on the graph
26
Differentiate endpoint and kinetic (multiple point/rate/continuous monitoring) enzyme methods, including why kinetic methods are preferred to endpoint methods for enzyme assays
Substrate depletion is observable telling us to dilute the sample
27
Michaelis-Menten curve:
- Zero Order kinetics
- Where on curve
At the end of the curve when it starts to flatten out
28
Michaelis-Menten curve:
- First Order kinetics
- Where on curve
At the beginning of the curve were it is increasing
29
Michaelis-Menten curve:
| How are the axis labeled?
X: [S], concentration of substrate (mol/L)
Y: V, initial reaction rate (mol/Ls)
30
Michaelis-Menten curve:
| Vmax and Km on the curve
Vmax: Highest point curve goes
Km: half way up the Vmax
31
6 factors that influence enzymatic activity
1. enzyme concentration
2. substrate concentration
3. temperature
4. pH
5. presence of cofactors
6. presence of inhibitors
32
Define plasma-specific enzymes
Expected to be in higher concentration in blood b/c they function there (i.e., coag factors--fibrinogen, thrombin)
33
Define non-plasma specific enzymes
Have no function in the plasma; function in tissues (cellular metabolism, etc.) (e.g., LD or CK)
34
Two GENERAL mechanisms for increased enzymes
1. increase in rate of enzyme being released into bloodstream
2. increase in rate of production of enzyme
35
7 specific causes of cell damage or death
1. hypoxia
2. chemicals and drugs
3. physical agents
4. microbiological agents
5. immune mechanisms
6. genetic defects
7. nutritional disorders
36
2 specific causes of increased enzyme production
1. Enzyme induction (via drugs, alcohol, etc)
| 2. Proliferation of cells that produce that enzyme (cancer)
37
List 5 physiological factors which affect enzyme reference ranges
1. Sampling time
2. age
3. sex
4. race
5. exercise (lack of and excessive)
38
Cholinesterase (ChE)
| - In vivo reaction that is catalyzed
Hydrolysis of choline esters to form choline and the corresponding fatty acid at the neuromuscular junction
39
What are the 5 major liver enzymes?
1. aspartate aminotransferase (AST)
2. alanine aminotransferase (ALT)
3. Alkaline phosphatase (ALP)
4. Gamma-glutamyltransferase (GGT)
5. Cholinesterase (ChE)
40
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Aspartate Aminotransferase (AST)
- Three biological sources
```
1. Heart
2. Skeletal muscle
3. Liver
(4. Kidney)
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Aspartate Aminotransferase (AST)
- typical appearance, peak, and return to normal after myocardial infarction
```
Rises w/in 12 hours after onset of chest pain; peaks at 18-24 hours; normal w/in 4-5 days
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Aspartate Aminotransferse (AST)
- Two liver diseases that give rise to the greatest elevations
```
Liver (hepatocellular) Disease
1. Viral hepatitis
2. Liver carcinoma
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Aspartate Aminotransferase (AST)
- One muscular disease in which AST is elevated
```
Skeletal muscle disease (Muscular Dystrophy)
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Aspartate Aminotransferase (AST)
- If hemolyzed specimens are unacceptable
```
UNACCEPTABLE due to high intracellular concentration of AST
45
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Alanine Aminotransferase (ALT)
- principle biological source
```
Liver
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Alanine Aminotransferase (ALT)
- Relative use in the dianosis of liver disease, compared to AST
```
Useful in the diagnosis of acute and chronic liver disease; parallels the rise in AST activity
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Alanine Aminotransferase (ALT)
- Liver disease in which ALT is elevated
```
Hepatitis; ALT is higher and persists longer than AST
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Alanine Aminotransferase (ALT)
- If hemolyzed specimen is unacceptable
```
NO HEMOLYSIS
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Alkaline Phosphatase (ALP)
- Four principle biological sources of total ALP
```
1. Liver
2. Bone
3. Intestine
4. Placenta
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Alanine Aminotransferase (ALT)
- Two liver diseases that gives rise to the greatest elevations
```
1. Hepatitis
| 2. hepatocellular carcinoma
51
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Alkaline Phosphatase (ALP)
- Hepatobiliary diseases in which ALP is increased
```
1. Biliary obstruction
2. Hepatitis
3. Cirrhosis
4. Infectious mono
5. Metastic carcinoma
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Alkaline Phosphatase (ALP)
- Hepatobiliary disease which gives rise to ALP's highest elevations
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Biliary obstruction
53
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Alkaline Phosphatase (ALP)
- Bone diseases in which ALP is increased
```
1. Bone tumors
2. Paget's disease
3. Rickets
4. Osteomalacia
5. Hyperparathyroidism
6. Healing fractures
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Alkaline Phosphatase (ALP)
- Two bone diseases which give rise to ALP's highest elevations
```
1. Bone tumors
| 2. Paget's disease
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Alkaline Phosphatase (ALP)
- If hemolyzed specimens are unacceptable
```
Gross hemolysis NOT acceptable (slight is ok, but must be documented)
56
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Alkaline Phosphatase (ALP)
- Principle of the Bowers and McComb procedure using p-Nitrophenylphosphate
```
Reference method; kinetic method that uses pNPP as a substrate and measures the rate of p-Nitrophenoxide release in AMP buffer
57
Which specific isoenzymes may be distinguished using the heat stability, urea denaturation, amino acid inhibition, and electrophoretic techniques?
Alkaline Phosphatase (ALP)
58
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Alkaline Phosphatase (ALP)
- Why is the reference range higher in children than in adults?
```
Due to rapid bone growth
59
Gamma-glutamyltransferase (GGT)
| - Two principle biological sources
1. Liver mainly
| 2. Kidney
60
Gamma-glutamyltransferase (GGT)
| - clinical usefulness in the diagnosis and monitoring of chronic alcoholism
GGT is induced by alcohol and other drugs; used as a marker to check for abstinence from alcohol
61
Cholinesterase (ChE)
| - Three clinical applications of measuring decreased ChE activity
1. Assess exposure to organophosphates found in insecticides and nerve gas
2. Check how patient will react to general anesthesia
3. Assess presence of cirrhosis, hepatitis, liver carcinoma (due to decreased production)
62
Acetylcholinesterase (AChE)
| - Biological sources
1. RBCs
| 2. CNS
63
Cholinesterase (ChE)
| - Biological sources
1. Liver
2. White matter of brain
3. Serum
64
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Creatine Kinase (CK)
- In vivo reaction it catalyzes
```
Catalysis for ATP formation and phosphorylation of creatinine in energy production or usage
65
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Creatine Kinase (CK)
- Three principle tissue sources
```
1. Skeletal muscle
2. Cardiac muscle
3. Brain
66
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Creatine Kinase (CK)
- dimeric composition and sources of its three isoenzymes
```
Composed of two subunits (M and B)
| Isoenzymes formed: CKMM, CKMB, CKBB
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Creatine Kinase (CK)
- Typical appearance, peak, and return to normal after a myocardial infarction
```
Rises 3-8 hours after the onset of chest pain (1st to rise)
Activity peaks at 10-36 hours
Levels return to normal in 3-4 days
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Creatine Kinase (CK)
- One muscular disease that gives rise to CK's highest elevations
```
Duchenne's Muscular Dystrophy (CKMM)
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Creatine Kinase (CK)
- Cerebral diseases in which CK is elevated
```
Cerebral vascular accident (CVA), stroke, verebral ischemia (CKBB)
70
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Creatine Kinase (CK)
- Principle of the Oliver and Rosalki method
```
Analytical procedure for total CK; increase in absorbance as NADP+ is oxidized and is measured at 340nm
71
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Creatine Kinase (CK)
- If hemolyzed specimens are acceptable
```
ACCEPTABLE
72
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Lactate Dehydrogenase (LDH)
- In vivo reaction it catalyzes
```
Catalyzes the interconversion of pyruvate and lactate in glycolytic pathway
73
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Lactate Dehydrogenase (LDH)
- Seven principle biological sources
```
1. Brain
2. RBCs
3. WBCs
4. Kidney
5. Liver
6. Lung
7. Cardiac muscle
8. Skeletal muscle
74
```
Lactate Dehydrogenase (LDH)
- Typical appearance, peak, and return to normal after a myocardial infarction
```
Rises 8-18 hours after onset of chest pain
Peaks ~48-72 hours
Levels return to normal in 6-10 days
75
```
Lactate Dehydrogenase (LDH)
- Liver diseases in which LDH is elevated
```
1. Obstructive jaundice
2. Cirrhosis
3. Viral hepatitis
4. infectious mononucleosis
5. Toxic jaundice
76
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Lactate Dehydrogenase (LDH)
- Liver disease that gives rise to LDH's highest elevation
```
Hepatobiliary disease (metastatic cancer)
77
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Lactate Dehydrogenase (LDH)
- Two types of anemias in which LDH is elevated
```
1. Megaloblastic anemia (can't absorb vitamin B12)
| 2. Pernicious anemia
78
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Lactate Dehydrogenase (LDH)
- If hemolyzed specimen is unacceptable
```
Acceptable, but make a comment. Hemolysis increases LDH in vivo or in vitro
79
Aldolase (ALS)
| - Three principle biological sources
1. Skeletal muscle
2. Liver
3. Brain
80
Aldolase (ALS)
| - One disease state in which ALS is elevated
Muscle dystrophy; can measure along w/ CK, which will also be greatly elevated
81
Amylase (AMS)
| - In vivo reaction it catalyzes
Breakdown of starch into amylose
82
Amylase (AMS)
| - Two principle biological sources
1. Pancreas (40% of total AMS)
| 2. Salivary glands (60% of total AMS)
83
Amylase (AMS)
| -Clinical significance of macroamylase
AMS is bound to IgG or IgA, causing an increase in total AMS w/o apparent disease
84
Amylase (AMS)
| - Clinical usefulness of AMS measurements in the diagnosis of acute pancreatitis, chronic pancreatitis, and the mumps
```
Acute pancreatitis
- Increased in serum shortly after onset (may be detectable in urine)
- Peaks w/in 24 hours
- Returns to normal w/in 4 days
Chronic pancreatitis
- ??
Mumps
- AMS is increased
```
85
Amylase (AMS)
| - What method measures amount of sugars formed from hydrolytic activity of AMS
Saccharogenic
86
Lipase (LPS)
| - Two diseases in which LPS is elevated
Acute and chronic pancreatitis
87
Lipase (LPS)
| - A comparison b/w the usefulness of LPS and AMS measurements in the diagnosis of pancreatitis
Levels parallel AMS appearance, but LPS is more specific than AMS
88
Lipase (LPS)
| - Principles of turbidimetric/nephelometric procedures
Olive oil emulsion is hydrolyzed by LPS, causing a DECREASE in turbidimetry
89
Lipase (LPS)
| - Principle of coupled enzymatic analytical procedure
LPS hydrolyzes fatty acids to form free glycerol which is quantitated colorimetrically
90
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Acid Phosphatase (ACP)
- Clinical usefulness of performing ACP measurements
```
Used to diagnose diseases of the prostate; can also do a prostatic acid phosphatase (PAP) and PSA
91
Glucose-6-phosphate dehydrogenase (G-6-PD)
| - Biological sources
RBCs, adrenal cortex, LN, thymus, spleen
92
Glucose-6-phosphate dehydrogenase (G-6-PD)
| -Disease state associated w/ LOW levels of G-6-PD
Hemolytic anemia (low levels of NADPH and glutathione which destroys Hgb causing RBCs to lyse)
93
Four major coenzymes used in electron transfer reactions commonly employed in enzyme assays in the lab, including specific wavelength for measurement
NAD+, NADH, NADP+, NADPH @ 340nm
94
Clinically significant enzymes/lab findings
| - Myocardial infarction
Increase in: CKMB, AST, LDH
95
Clinically significant enzymes/lab findings
| - Liver (hepatocellular) disease
AST, ALT
96
Clinically significant enzymes/lab findings
| - Liver (hepatobiliary) disease
GGT, ALP
97
Clinically significant enzymes/lab findings
| - Bone disease
ALP
98
Clinically significant enzymes/lab findings
| - Brain disease
CKBB
99
Clinically significant enzymes/lab findings
| - Prostate cancer and hypertrophy
ACP
100
Clinically significant enzymes/lab findings
| - Acute and chronic pancreatitis
LPS, AMS
101
Clinically significant enzymes/lab findings
| - Muscle disease and muscular dystrophy
CKMM, ALS
102
Amylase (AMS)
| - What method uses dye-labeled starch substrates; as AMS hydrolyzes the starch, an increase in color is quantified
Chromilytic
103
Amylase (AMS)
| - What method measures the decrease in starch substrate concentration as AMS works on starch?
Amyloclastic assays
104
Amylase (AMS)
| - What method consists of a glucose coupled reaction and a hexokinase coupled reaction?
Enzymatic assays
