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59 Cards in this Set
- Front
- Back
- 3rd side (hint)
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Enzyme inhibition
How many forms? |
2
List |
Reversible Irreversible |
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Enzyme inhibition
Reversible _______ |
Irreversible
Mechanism |
Covalent bond (sharing of e-) between E and S |
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Irreversible enzyme inhibition
Enzyme activity cannot be restored with _________ |
Removal of inhibitor
Can it reverse the irreversible enzyme inhibition? |
N |
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irreversible enzyme inhibition
Enzyme activity cannot be _____ with removal of inhibitor. |
_______ cannot be restored with _____ of inhibitor |
Enzyme activity removal |
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Irreversible enzyme inhibition
Enzyme activity cannot be restored with removal of inhibitor
Notes |
Chemical warfare agents (nerve gas)
Mechanism |
Inhibits acetylcholinesterase |
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Enzyme inhibition
2 forms |
_________+ how many different forms there are? Irreversible |
Reversible
3 |
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Reversible enzyme inhibition
3 forms |
Competitive Noncompetitive Uncompetitive |
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Reversible enzyme inhibition
3 forms
_______ Noncompetitive Uncompetitive |
Competitive
Definition |
The substance is similar to the normal substrate and competes with the substrate for the binding or active site of the enzyme. |
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Reversible enzyme inhibition
Competitive ______________ Uncompetitive |
Noncompetitive
Definition |
The inhibitor is structurally different from the substrate and Binds to an allosteric site of the enzyme or ES |
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Competitive inhibitor
Structure |
Similar to the substrate
where does it bind to? |
Bind to the same binding or active site of the enzyme |
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Noncompetitive inhibitor
Structure |
Structurally different from the substrate
Where does it bind to? |
Allosteric site on enzyme or ES |
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Noncompetitive inhibitor
aka |
Mixed inhibition
Structure Where does it bind to? |
Structurally different from substrate
Bind to allosteric site of enzyme or ES |
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Reversible enzyme inhibition
3 forms Competitive Noncompetitive ___________ |
Uncompetitive inhibition
Definition |
Inhibitor binds to the ES complex to form an enzyme-substrate inhibiting complex that does not yield product |
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Uncompetitive inhibition
Where does the inhibitor bind to? |
ES complex
Result |
ES inhibiting complex |
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Uncompetitive inhibition
Inhibitor binds to ES complex to form ES-inhibiting complex
End result? |
No product |
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Does uncompetitive inhibition yield products? |
No
Where does it bind to? |
ES complex |
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V vs. S graph
No inhibitor vs. Competitive inhibition |
Same V max
line is lower
Formula and conclusion |
E+S (K1/K-1) ES (K2) E+P
EI+S: no reaction |
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V vs. S graph
No inhibitor vs. Noncompetitive inhibition |
Lowered V max Line is lowered
Formula and conclusion |
E+S (K1/K-1) ES (K2) E+P
Some products formed, but reduced |
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V vs. S graph
No inhibitor vs. Uncompetitive inhibition
Formula |
E+S (K1/K-1) ES (K2) E+P ESI
Conclusion |
no reaction |
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Lineweaver-Burk plot
m: (Slope) |
Km/Vmax
x |
1/S |
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Lineweaver-Burk plot
b |
1/Vmax
m? |
km/Vmax |
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Lineweaver-Burk plot
X axis Closer to 0/Y |
High conc.
Away from 0/Y |
Lower conc. |
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Lineweaver-Burk plot
Y axis Closer to 0/x |
Faster
Away from 0/x |
Slower |
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Lineweaver-Burk plot
No inhibition vs. Competitive inhibition
What is unchanged? |
1/Vmax
How to achieve the same V max? |
Add more S |
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Lineweaver-Burk plot
No inhibition vs. Competitive inhibition
What is changed? |
Decreased 1/Km in the inhibited rxn
Why? |
Km is a fixed value for each enzyme
Recall slope is Km/Vmax |
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Lineweaver-Burk plot
Competitive inhibition
Higher conc. or closer to Y/0 |
Faster
Away from Y/0 |
Slower |
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Lineweaver-burk plot
Non-competitive inhibition
what is unchanged? |
-1/Km
Regular graph What is unchanged? |
Km |
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Lineweaver-Burk plot
Why -1/Km is unchanged in non-competitive inhibition? |
+ S will not influence velocity |
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Non-competitive inhibition
Lineweaver-Burk plot
What is changed? |
1/Vmax
Regular graph What is changed? |
V max |
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Lineweaver-Burk plot
How 1/Vmax is changed? |
Increased
Why it is increased? |
The inhibitor slows the rxn original Vmax can't be achieved |
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Non-competitive inhibition
Reversible? |
no
Why? |
The binding of inhibitor is independent of the substrate. |
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Non-competitive inhibition
1/Vmax is increased
meaning |
Rxn is slower |
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Non-competitive inhibition is nonreversible because? |
Binding of inhibitor is independent of the substrate
Adding more substrate? |
Will not remove inhibition |
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Uncompetitive inhibition
Original graph
What is changed? |
Both Km and V max
How they are changed? |
both are decreased |
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Uncompetitive inhibition
Lineweaver-Burk plot
What is changed? |
Both -1/Km and 1/Vmax
How they are changed? |
-1/Km is increased
1/Vmax is also increased |
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Uncompetitive inhibition
1/Vmax is increased?
Meaning |
Slower rxn |
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Uncompetitive inhibition
occurs when the inhibitor binds to the ES complex
First require ES formation
This causes an apparent decrease in Km
Notes |
Km is ______ for an enzyme |
fixed |
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_____ is fixed for an enzyme |
Km is fixed for an ______ |
enzyme |
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How sensitive enzymes are to Temperature? |
Extremely sensitive
Generally, every 10 degree increase in temperature will result in? |
doubled enzyme activity |
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Doubled enzyme activity
Temperature |
+10 C
but the actual difference is dependent on? |
enzyme |
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Enzymes' optimal temperature |
Body temperature |
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Effects of high temperature on enzymes |
Denaturation
Examples Optimal temperature for
1. CK 2. Amylase |
1. 10 2. 45 C |
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high temperature leads to denaturation of enzymes
Exceptions? |
Taq polymerase (PCR) stable at high temperature
How high? |
95C |
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Optimal pH for each enzymes is dependent on? |
Active site's specific ionization
Note |
best to choose pH at this optimal level |
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range of optimal pH
different enzymes |
some have border pH range, meaning? |
Less critical |
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Some enzymes have broader pH range, so this is less critical
Example |
ALP
Max activity pH |
9-10 |
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Isoenzymes and pH values |
They often have different pH values
Solution |
Compromise to get activity of all isoenzymes at once. |
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Isoenzymes often have different pH values, so compromise is made to get activity of all isoenzymes at once
Alternatives |
Immunoassays
Application |
Measure the mass of enzyme protein |
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How can ionic strength of solution affects enzyme activity? |
If too high, activity goes down
Example |
NaCl |
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If we dilute with saline (NaCl), what could happen to enzyme actiivty |
It could adversely affect enzyme activity.
Enzyme activity if too high? |
Activity goes down. |
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Proteins and enzyme activity |
proteins are needed to maintain enzyme activity
Mechanism |
Preventing denaturation |
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Proteins are needed to maintain enzyme activity by preventing denaturation
When patient sample out of linear range, solution? |
Enzyme diluent containing _________________ is needed |
plasma proteins |
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When patient sample is out of linear range, enzyme diluent containing plasma proteins is needed
Example |
Plasma
Urine |
70 g/L
0 |
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Albumin and enzyme activity |
It will increase activity of urinary amylase. |
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Cofactors or coenzymes vs. rate of rxn |
+ |
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Cofactors
Metal activators
Example |
Mg++
Mechanism |
attract - charged group |
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Cofactors
Coenzymes
Mechanism |
e- donors and acceptors
Example |
NAD+ NADH |
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Coenzymes are e- donors and acceptors
How they are added? |
Add these activated coenzymes to the enzyme reagent in excess. |
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Lipase measured with colipase with cofactor |
5-10 x faster |
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