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9. Enzyme kinetics


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What are the 4 physical properties that we identify in enzymes?
1. Composition
2. Cofactors used
3. Isoenzyme forms
4. Binding sites
What are enzymes composed of?
Proteins, organic components of C, H, O, N, S,.
What is a catalyst?
Substance that increases a reaction rate without being changed or consumed.
What are cofactors?
Nonproteins that optimize an enzyme's catalytic activity.
What is a
Holo = whole enzyme+cofactor
Apo = enzyme w/out cofactor
What are
-Prosthetic groups?
Coenzyme: loose bound organic cofactor.
Activator: loose inorganic ion
Prosth. Grp: titely bound cofactor
What are characteristics of isoenzymes?
Different forms that catalyze same reaction; unstable; differentiate w/ electrophoresis
-CK and LD are examples
What are the catalytic properties of enzymes?
1. Effective in small amounts
2. Unchanged by the reaction
3. Decrease activation energy
4. Specific
5. Can lose enzyme activity
What causes an enzyme to irreversibly lose its activity?
4 types of enzyme specificity:
1. Absolute
2. Group
3. Bond
4. Stereo
What are the 2 types of enzyme classifciations?
1. Substrate based
2. IUB systematic
How are enzymes classified in the old Substrate based method?
Based on the type of SUBSTRATE the enzyme acts upon
How are enzymes classified in the new IUB systematic method?
Based on the catalyzed REACTION
List the 6 enzyme classes in the IUB systematic method:
1. Oxidoreductases
2. Transferases
3. Hydrolases
4. Lyases
5. Isomerases
6. Ligases
-Cite an oxidoreductase:
-Cite 4 transferases:
-Cite 3 hydrolases
Oxidored: LD
Transfer: CK, AST, ALT, GGT
Hydrolas: Acid phosphatase, amylase, urease
What are the 3 phases of an enzyme reaction?
1. Lag phase
2. Linear phase
3. Substrate depletion phase
What occurs in the lag phase?
Mixing and temp/kinetic equilibrium establishment.
What occurs in the linear phase?
Constant reaction; the change in absorbance is constant per unit time - this is where we assay.
What occurs in the substrate depletion phase?
little change in absorbance/time
What substrate concentration is seen in 1st vs. Zero order reactions?
How does the reaction rate relate to each order?
1st order: low [S] - rate is proportional to amt of substrate

Zero order: high [S] - rate is independent of substrate
Why is zero order reaction rate preferred to first order?
Because the zero order reaction will produce a linear graph of the reaction and allow measurement of the enzyme.
What is the definition of the michaelis menton constant?
The [S] that gives 1/2Vmax.
What is the substrate concentration used in clinical assays?
How does that Substrate conc. affect the Velocity? What order is the reaction? What happens to the Vmax?
-Velocity approaches Vmax.
-Order is Zero
-Vmax becomes indepenent of [S]
So what is the resulting rate of clinical, zero-order assays?
>90% Vmax!
What is the optimum pH for biologic reactions?
What temp are clinical enzymes measured at?
Biological, 37
On a lineweaver-burk plot:
-What is the y-intercept?
-What is the slope?
Y-intercept = 1/Vmax

Slope = Km/Vmax
So if a Km is increased but Vmax is unchanged, what happens to the plot?
-What type of inhibition is this?
The slope increases but the y-intercept stays the same.
-Competitive inhibition
If the Km is unchanged but the Vmax is decreased what happens to the plot?
-What type of inhibition is this?
Slope increases AND the y-intercept increases.
-Noncompetitive inhibition
What happens to a lineweaver burke plot when Vmax AND Km are decreased?
-What type of inhibition is this?
The Y-intercept decreases but the slope stays the same.
-Uncompetitive inhibition

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