Introduction to Enzyme Kinetics Flashcards
How do enzymes speed up the rate of reactions?
By lowering the free energy activation for a reaction. They do not change the standard free energy of the product
First order reaction
- Reaction rate is directly proportional to substrate concentration
- Enzyme kinetics follow this pattern prior to reaching Vmax
Zero order reaction
- The maximum velocity of a reaction is reached and active sites are almost continuously filled
- Increase in substrate concentration after this point will not increase the rate
Michaelis constant (Km)
Substrate concentration required to reach one half of Vmax
At this [S], half of the enzymes are bound to S
How are Km and Enzyme/Substrate affinity related?
They are inversily related
Higher Km = lower affinity
Lower Km = higher affinity
The enzyme velocity is at one-half the max rate when what percent of the enzyme is bound to substrate?
50% of the enzyme is bound to substrate
What happens to the Vmax when more enzyme is added?
The Vmax increases
What happens to the Vmax when the enzyme concentration is reduced by 50%?
The Vmax is reduced by 50%
What does Kcat represent?
The turnover number which describes how many substrate molecules are transformed into products per unit time by a single enzyme
Lineweaver-Burk plot
- X-axis = 1/[S]
- X-intercept = 1/Km
- Y-axis = 1/Vo
- Y-intercept = 1/Vmax
- Slope = Km/Vmax
What are the 4 main types of enzyme inhibition?
- Competitive inhibition
- Non-competitive inhibition
- Irreversible inhibition
- Allosteric enzyme
Competitive inhibition
- Competes with a substrate for binding at the enzyme’s substrate-recognition site
- Vmax = unchanged
- Competitive inhibitors can be overcome by increasing the substrate concentration
- Km = increased
- More substrate is needed to achhieve 1/2 Vmax
Competitive inhibition on plots
Noncompetitive inhibition
- Inhibitor and substrate bind at different sites
- Inhibitor can bind E or ES complex and prevents the reaction from occuring
- Vmax = Decreases Vmax
- Cannot be overcome with additional substrate
- Km = unchanged
- Noncompetitive inhibitors do not interfere with the binding of substrate thus do not change the affinity
Noncompetitive inhibition plots
Irreversible inhibition
- Structural analog of the substrate is converted to a more effective inhibitor with the help of the enzyme to be inhibited
- New product irreversibly binds to the enzyme and inhibits further reaction
- If [inhibitor] > [enzyme] –> No reactions as all enzymes are inhibited
What are three main factors that influence enzymatic activity?
-
Temperature: rate of reaction increases with temperature then decreases as enzymes denature at high temperatures
- Most animal enzymes rapidly denature above 40 C
-
pH: Different enzymes have different optimal pHs
- Some enzymes are only function at high or low pHs
- Substrate concentration: increases in [S] increase velocity up to the point of Vmax at which point more S no longer increases the velocity
Allosteric enzymes
- Regulated by molecules called effectors that noncovalently bind at a site other than the active site
- Composed of multiple subunits and regulatory (allosteric) site that bind effector
- Frequently catalyze commited step early in a pathway
Effector
Molecule that regulates allosteric enzymes by noncovalently binding at a site other than the active site
Positive effector
Either increases the Vmax or decreases the Km
Negative effector
Either decreases Vmax or increases Km
Homotropic effector
- When the substrate iteself serves as an effector
- Often (but not always) are positive effectors, where the presence of substrate molecule at one site enhances the catalytic properties of the other site
- This is called cooperativity –> Exhibits a sigmoidal curve
Heterotropic effector
The effector is different from the substrate
What type of allosteric effect does carbon dioxide have on hemoglobin’s ability to carry oxygen?
CO2 acts as a negative, heterotropic, allosteric effector
What step are regulatory enzymes usually involved in?
The rate-limiting step or the commited step
