Enzymes Flashcards
State the definition and five general properties of enzymes, according to the lecturer.
- Enzymes are 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
- Each enzyme is highly specific for a given reaction
- Enzymes act by lowering the activation energy
Recognize examples of activators (inorganic cofactors) and coenzymes (organic cofactors).
a. Activators (inorganic cofactors): Zn, Fe, Cu, Mg, Mn
b. Coenzymes (organic cofactors): NAD, NADH, NADP, NADPH, Pyrixodal-5-phosphate
Explain why enzyme activity, rather than enzyme concentration, is measured in the laboratory.
The amount of enzyme is so small that it’s difficult to devise assays that are sensitive enough
The rate of product formation/amount of substrate consumed during an enzymatic reaction is measured
What is occurring during the lag, log, and substrate depletion phases
a. Lag phase – initial period when enzyme and substrate (specimen) are first mixed, but no product is formed – no change in absorbance
b. Log (linear) phase – rate of product formation is linear with time
c. Substrate depletion phase – No more substrate remaining to convert to product – no change in absorbance – rate of reaction is proportional to substrate concentration
Specifically why enzyme measurements should be made during the log phase, according to the principles of Beer’s Law
The amount of product being produced is a function of enzyme concentration
d. Definitions of zero order and first order kinetics
a. Zero order: the rate of the reaction is independent of the substrate (reactant)
b. First order: the rate of the reaction is proportional to the substrate (reactant)
- Differentiate endpoint and kinetic (multiple point/rate/continuous monitoring) enzyme methods, including why kinetic methods are preferred to endpoint methods for enzyme assays.
a. Endpoint method: the measured product is no longer changing because ALL of the analyte has reacted with the reagent in the sample – the absorbance remains fixed
b. Enzymatic Rate (Continuous Monitoring): the difference in absorbance between two (2) points during the progress of a reaction – allows for measurements to be taken while the enzyme is still reacting
Six factors that influence enzymatic activity
- Enzyme Concentration
- Substrate Concentration
- Temperature
- pH
- Presence of cofactors
- Presence of inhibitors
Recognize the importance of maintaining proper temperature and pH reaction conditions while performing enzyme assays.
The closer the assay temperature is to the normal body temp (37 C), the faster the reaction, but denaturation will occur at temperatures higher and lower than that – must be closely monitored
Buffers are incorporated into assays to maintain optimal pH and eliminate denaturation
Differentiate plasma-specific and non-plasma-specific enzymes in detail according to:
a. Definition
b. Expected relative plasma and tissue concentrations
a. Plasma-specific enzymes: known function in the plasma (i.e. coagulation factors)
Non-plasma specific enzymes: no known function in the plasma
b. Plasma-specific enzymes: higher concentration in blood relative to the tissue
Non-plasma specific enzymes: higher concentration in tissue relative to the blood
Discuss the two mechanisms for an increased rate of entry of enzymes into circulation in detail, according to:
a. Two general processes involved
b. At least two specific causes of cell damage or death
c. Two specific causes of increased enzyme production
a. Increase in the rate of enzyme release into the bloodstream
Increase in the rate of production of an enzyme
b. Hypoxia
Chemicals and drugs Physical agents (i.e. trauma)
c. Enzyme induction (via drugs, alcohol)
Proliferation of cells (that produce the enzyme -- cancer)
Recognize five physiological factors which affect enzyme reference ranges.
- Sampling time (i.e. circadian rhythm time after onset of chest pain)
- Age
- Sex
- Race
- Exercise
Discuss aspartate aminotransferase (AST) according to:
a. Three biological sources
b. Two liver diseases that gives rise to the greatest elevations
c. One muscular disease in which AST is elevated
d. One cardiac disease in which AST is elevated
e. If hemolyzed specimens are unacceptable
a. Cardiac muscle
Skeletal muscle
Liver
b. Viral hepatitis
Liver carcinoma
(Cirrhosis)
c. Muscular dystrophy
d. Myocardial Infarction: rises with 12 hrs. of onset, peaks at 18-24 hrs., normal within 4-5 days
e. YES
- Discuss alanine aminotransferase (ALT) according to:
a. Principal biological source
b. Relative use in the diagnosis of liver disease, compared to AST
c. Liver diseases in which it is elevated
d. Two liver diseases that gives rise to the greatest elevations
e. If hemolyzed specimens are unacceptable
a. Liver
b. Parallels the rise in AST activity
usually ALT > AST (persists longer as well)
AST > ALT in cirrhosis
c. Hepatic necrosis
Cirrhosis
d. Viral hepatitis
Liver carcinoma
e. YES
State the condition in which AST levels are expected to be greater than ALT levels.
Cirrhosis
