Unit 5 - Enzymes Flashcards
What structure are enzymes in to be functional?
Quaternary
What are enzymes composed of?
Subunits or protomers grouped together
How many subunits do enzymes need to be functional?
All of them
Isoenzymes
Different forms of enzymes that catalyze the same reaction
Subunits
AKA promoters
Grouped together to form a fully functional enzyme
Function of enzymes
Increase rate of a reaction
Enzymes only increase the rate of a reaction not…
create a reaction that wouldn’t normally occur
How do enzymes increase rate of reaction?
Lower activation energy
Where do enzymes get synthesized?
Intracellularly
Specific enzyme synthesis markers
PSA - prostate only
CK - heart and skeletal muscle
Wide variety synthesis markers
LD - Different tissues
ALP - Bone Liver Intestine Placenta (BLIP)
Why is half life important in measuring enzymes?
You know if cell damage was recent or not
Active site
Catalytic site of enzyme
Where do substrates bind
Active site
Law of mass action
Direction of reaction from high concentration to low concentration
Enzyme cofactors bind what
Bind allosteric site
What do enzymes need to work?
Cofactors
Apoenzyme
Protein part of enzyme WITHOUT cofactor
Holoenzyme
Functional enzyme with cofactors/all parts
Are cofactors consumed after the reaction?
No
Types of cofactors
Activators
Coenzymes
Prosthetic groups
Activator cofactors
Inorganic ions like ca or mg
Coenzyme cofactor
Non protein organic compound (NAD/NADH)
Prosthetic group cofactor
Cofactor bound so tightly it looks like its part of the enzyme
Stereoisomeric specificity
Enzymes that only work with a specific isomer (3d config)
Isomer
3D configuration
When would enzymes release?
When there’s damage to cells
Enzyme activity
Amount of substrate converted to a product in a given period of time
Standard International Unit of Enzyme Activity
Quantity of enzyme that catalyzes a rxn of one micromole of substrate per minute
Enzyme equilibrium (Michaelis-Menten)
Point where product is formed from substrate at a constant RATE
Rate =
Velocity under constant conditions
Constant rate of the enzyme run is proportional to
Amount of Substrate available
Amount of Enzyme available
How much enzyme-substrate complex made
How much product made
What is the rate limiting factors of an enzyme reaction
Substrate
Km
M-M constant (1/2 Vmax)
Substrate concentration at which enzyme catalyzes 1/2 as fast as it can
Low substrate concentration
Enzyme makes product slowly
First order kinetics
Product formation depends on substrate concentration
(low substrate)
High Substrate concentration
Enzyme makes product faster
Vmax
Maximum velocity
Zero order kinetics
Substrate in excess relative to enzyme
(high substrate concentration)
Mixed order kinetics
Center of the graph where its not predictable
Which part of the MM curve shows zero order kinetics
The top where Vmax is
Which part of the MM curve shows first order kinetics
The bottom where the initial velocity is slow because there less substrate
When Km is determined, what do you have to do with the conditions that achieved that?
Keep them the same, they have to be exactly duplicated
What should be in excess for expected enzyme concentration?
Substrate
What should be in excess for expected substrate concentration?
Enzyme
What does the Km have to be multiplied by for enzyme concentration?
Multiply Km by 10-100 fold
What does the Km have to be substrate concentration?
Half of Km
What does it mean if one substrate has a higher Km than the other?
Low affinity, requires more enzyme to do the job
What does it mean if one substrate has a lower Km than the other?
High affinity, requires less enzyme to do the job
Do you want a substrate with a low Km or high Km?
Low Km
International standard temp
30C
Frequently used temp for enzymes
37C
Lag period
Little product formed, reaction getting started
Linear period
Equilibrium attained
Product formed at a constant rate (Vmax)
Nonlinear period
No additional product formed, substrate exhaustion
The more product formed per minute…
The more enzyme activity, the more enzyme concentration
When using enzyme to measure substrate, what do you want in excess?
Enzyme or co-factors
When using enzyme to measure substrate, what controls the rate of the reaction
Substrate
When using enzyme to measure substrate, what order kinetics must be observed?
First, you want low substrate to find the linearity point
What is on the x and y axis of a line weaver burk plot
Y: 1/Vmax
X: -1/Km
Which plot is used to study inhibitor activity?
Lineweaver burk plot
If the X intercept moves to the left, what does that mean for the Km?
Km lower
Better enzyme activity
If the X intercept moves to the right, what does that mean for the Km?
Km is higher
Worse enzyme activity
If the Y intercept moves up, what does that mean for the reaction?
Slowing down
IF the Y intercept moves down, what does that mean for the reaction?
Going faster
How do competitive inhibitors affect Km?
Km increases because more normal substrate needed to perform the same amount of enzyme activity
How do competitive inhibitors affect Vmax?
Vmax not affected because enough normal substrate can overcome inhibitor because of the large concentration
moves to right and up
worse enzyme
moves left and down
better enzyme
How does noncompetitive inhibition affect Km
No effect because there are still normal enzymes that are doing the reaction at that linear speed
Where do noncompetitive inhibitors bind
Allosteric site instead of active site
How does noncompetitive inhibition affect Vmax
Decrease
More enzyme is required to produce a product because enzyme are having their conformation changed
What is uncompetitive inhibition and hwo does it affect Km/Vmax
Binds enzyme-substrate complex
affects both
