Chapter 2: Enzymes Flashcards
What is an ACE inhibitor?
Angiotensin-converting enzyme (ACE) catalyze a reaction that converts angiotensin I to angiotensin II. Angiotensin II peptide causes restriction of the blood vessels to raise blood pressure and stimulates the release of the hormone aldosterone which activates the kidneys to absorb more water back into the bloodstream.
The constriction of the blood vessels along with the additional volume of water increases blood pressure.
ACE inhibitors stop the pathway by inhibiting ACE, effectively dropping blood pressure.
What do enzymes do? Do enzymes impact the thermodynamics of a biological reaction? Do enzymes change during the course of a reaction?
Enzymes increase the rate of biological reactions.
Catalysts do not change the thermodynamics of a reaction as deltaHrxn and equilibrium position do not change.
Enzymes, as catalysts, do not change during the course of a reaction.
Do enzymes raise or lower energy of activation?
Do enzymes increase or decrease the rate of a reaction?
Do enzymes alter the equilibrium constant?
Are enzymes changed or consumed in the reaction?
Are enzymes pH and temperature sensitive?
Do enzymes affect the overall deltaG?
Are enzymes specific or general in nature?
They lower Ae.
They increase the rate of rxn.
They do not alter equilibrium constant.
They are not changed or consumed in the rxn (they appear in both the reactants and products)
They are pH and T sensitive and have optimal activity at specific pH and T.
They do not affect the overall deltaG.
Enzymes are considered highly specific for a particular reaction or class of reactions.
What are the six classifications of enzymes?
LIL HOT
Ligase
Isomerase
Lyase
Hydrolase
Oxireductase
Tranferase
What is a ligase? Do they usually require ATP? What kind of molecule (generally) do ligase act upon? How do they generally differ from lyases?
Ligase are enzymes that catalyze addition or synthesis reactions.
They generally occur between large similar molecules and often require ATP.
Synthesis of smaller molecules are generally accomplished by lyses.
What is an isomerase?
Isomerase are enzymes that catalyze the rearrangement of bonds within a molecule. Sometimes classified as oxireductases, transferases, or lyases, depending on the mechanism of the enzyme.
Isomerase catalyze reactions between stereoisomers as well as constitutional isomers.
What is a lyase? Do they require water as a substrate? Do they act as oxireductases? What is a synthase?
Lyase are enzymes that catalyze the cleavage of a single molecule into two products.
They do not require water or act as oxireductases.
Because most enzymes also catalyze the reverse of their specific reactions, the synthesis of the two molecules into a single molecule may also be catalyzed by a lyase, referred to as a synthase.
What is a hydrolase? How are many hydrolases named?
Hydrolases are enzymes that catalyze the breaking of a compound into two molecules using the addition of water.
Hydrolases are typically named for their substrate. Examples are:
phosphatase, cleave phosphate group from another molecule.
Peptidases, nucleases, lipases, breaking down proteins, nucleic acids, and lipids, respectively.
What are transferases? Are kinases transferases?
Transferases are enzymes that catalyze the movement of a functional group from one molecule to another.
Kinases are a member of class of transferases.
Remember serine/threonine protein kinases phosphorylate serine and threonine respectively.
What is a kinase?
A kinase is an enzyme classified as a transferase.
Kinases transfer a phosphate group, generally from ATP, to another molecule.
Again, what is a kinase.
Kinase are transferases that catalyze the transfer of a phosphate group, generally from ATP, to another molecule.
Serine/threonine kinase. Serine and threonine can be phosphorylated by serine/threonine kinase. These phosphorylated AA have important function throughout the body.
What is an oxidoreductase? Give a relevant example of a cofactor for oxidoreductases. (hint: oxidation/reduction of glucose)
How do the names of enzymes clue us to them being oxidoreductase?
An oxidoreductase is an enzyme that catalyzes oxidation-reduction reactions (the transfer of electrons between biological molecules).
Enzymes
NAD+ and NADH+ are cofactors that act as electron carries for reactions catalyzed by oxidoreductases.
Dehydrogenase or reductase in their names are usually oxidoreductases. Oxidases are enzymes that utilize oxygen as their final electron acceptor.
In an oxidoreductase reaction, what is the term for the electron donor? Acceptor?
In reactions catalyzed by oxidoreductases, the electron donor is known as the reductant, the electron acceptor is known as the oxidant.
What classification of enzyme is an oxidase?
An oxidase is an oxidoreductase enzyme that uses oxygen as the final electron acceptor.
What classification of enzyme is a dehydrogenase or reductase?
Dehydrogenase and reductase are usually oxidoreductases.
Reductant: electron donor
Oxidant: electron acceptor.
In oxidoreductases, what is a reductant? An oxidant?
Reductant donates electrons.
Oxidant accepts electrons.
Again. What are the major classification of enzymes?
LIL HOT
Lyase
Isomerase
Ligase
Hydrolase
Oxidoreductase
Transferase
What is an endergonic reaction? Exergonic?
Endergonic requires energy input (deltaG>0) Endo means in.
Exergonic energy is given off (deltaG<0) Exo means out.
Do enzymes change deltaG? What do they do?
Enzymes do not change deltaG of a rxn. Recall that they cannot turn an endergonic reaction into an exergonic reaction.
Enzymes reduce the activation energy required for a reaction, essentially making it easier for a substrate to reach the transition state.
MCAT concept check enzymes page 53 question 1
How do enzymes work as biological catalysts?
They reduce the activation energy of a reaction, thus speeding up the reaction and they are not used up in the reaction.
MCAT concept check enzymes page 53 question 2
What is enzyme specificity?
Enzyme specificity refers to the idea that a given enzyme will only catalyze a given reaction or type of reaction.
MCAT concept check enzymes page 53 question 3
What is the name for the molecule upon which an enzyme acts?
Substrate
What is the name for the interaction between an enzyme and substrate?
Enzyme substrate complex
What is the name for the site within and enzyme where a substrate is held?
Active site
Image of enzyme substrate binding
What is the lock and key theory for enzyme substrate binding?
What is the induced fit model of enzyme substrate binding?
What is a cofactor? Coenzyme?
They both act as activators of enzymes. They regulators induce a conformational change in the enzyme that promotes its activity.
Cofactors are generally inorganic molecules or metal ions and are often ingested as dietary minerals.
Coenzymes are small organic groups like vitamins or derivatives of vitamins such as NAD+, FAD, and coenzyme A.
What is a prosthetic group? Give an example.
Tightly bound cofactors or coenzymes that are necessary for enzyme function are known as prosthetic groups.
Heme group in hemoglobin is an examine of a prosthetic group. Heme is a porphyrin ring containing a ferrous iron ion (Fe2+).
MCAT concept check mechanism of enzyme activity page 57 question 1
How do the lock and key theory and induced fit model differ?
MCAT concept check mechanism of enzyme activity page 57 question 2
What do cofactors and coenzymes do? How do they differ?
They both act as activators of enzymes.
Cofactors tend to be inorganic (minerals), while coenzymes tend to be small organic compounds (vitamins).
They both induce conformational change in the enzyme thus promoted its activity.
Tightly bound cofactors or coenzymes that are necessary for enzyme function are termed prosthetic groups.
What are the fat soluble vitamins?
A, D, E, K
List of B vitamins
What is saturation regarding kinetics of monomer enzymes?
Saturation refers to the maximum capacity of a group of enzymes to act.
When the amount of enzyme is the limiting factor, the reaction will reach saturation where the enzymes are working at maximum velocity, called Vmax.
What is the only way to increase Vmax of an enzymatic reaction?
Increasing the concentration of enzyme is the only way to increase Vmax.
Draw a Michaelis Menten plot of enzyme kinetics.
As the amount of substrate increases, the enzyme is able to increase its rate if reaction until it reaches a maximum enzymatic reaction rate (Vmax). Once Vmax is reached, adding more substrate will not increase the rate of reaction.
Write an equation that relates the formation of product using an enzyme. (Enzyme, Substrate, k1, k-1, kcat, and Product)
What is the Michaelis Menten equation?
Under constant concentration of enzyme, we can calculate the velocity of the enzyme to substrate concentration using the Michaelis Menten equation.
Using the Michaelis Menten equation, when the reaction rate is equal to half, what does Km equal?
Km=concentration of substrate at 1/2(Vmax)
What is Km? What does a high Km mean? A low Km?
Km is a measure of affinity of the enzyme for its substrate.
Km is the Michaelis Menten constant and is often used to compare enzymes.
High Km means the enzyme has low affinity for its substrate.
Low Km means the enzyme has high affinity for its substrate.
What does a high Km mean? A low Km?
A low Km reflects a high affinity for the substrate (low [S] required for 50% of enzyme saturation)
A high Km reflects a low affinity for the substrate (high [S] required for 50% of enzyme saturation).
Is Km an intrinsic property of the enzyme substrate system?
Yes. It cannot be altered by changing the concentration of enzyme or substrate.
Recall that Km=[S] at 1/2(Vmax)
What is Kcat?
The variable Vmax represents maximum enzyme velocity and is measured in moles of enzyme per second. Vmax can be mathematically related to Kcat which has units of s^-1.
Kcat measures the number of substrate molecules “turned over” to product, per enzyme molecule per second.
What is the unit of Kcat? What is Kcat?
The units of Kcat are s-1. Kcat measures the number of substrate molecules converted to product per enzyme molecule per second.
What value Kcat do most enzymes have?
Most enzymes have Kcat values between 101 and 103.
Represent Vmax relating concentration of enzyme and Kcat.
Restate the Michaelis Menten equation using Kcat.
At very low [S], further simplify the Michaelis Menten equation.
What is a Lineweaver-Burk plot?
LW-B plots are handy because they give us an easier and more accurate approximation of Vmax. The M-M plot forces us to guess Vmax as it appears asymptotic.
LW-B is a double inverse plot of the Michaelis Menten equation.
Again. What is a Lineweaver-Burk plot?
Lineweaver burn plot is a double reciprocal plot of the Michaelis Menten equation. It allows us to determine the type of inhibition that an enzyme is experiencing because Vmax and Km can be compared without estimation.
Some enzymes don’t show the normal hyperbole when graphed on an M-M plot (v vs. [S]).
What shape do they show? Why? What example could we remember that exhibits a sigmoidal curve of v vs. [S]?
Some enzymes show a sigmoidal (S-shaped) kinetics owing to cooperativity among substrate binding sites.
The cooperative binding of hemoglobin, which acts as a transport protein rather than an enzyme, results in the characteristic sigmoidal curve.
What are cooperative enzymes? What are T and R states?
Cooperative enzymes have multiple subunits and multiple active sites.
Subunits and enzymes may exist in one of two states:
T : low affinity tense state
R : high affinity relaxed state
What causes cooperative enzymes transition from T to R state?
Binding of the substrate encourages the transition of other subunits from the T state to the R state, which increases the likelihood of substrate binding by these other sub units.
Conversely, loss of substrate can encourage the transition from the our state to the T state, and promote dissociation of a substrate from the remaining subunits.
How can we think cooperative enzymes like a party?
As more people start arriving, the atmosphere becomes more relaxed in the party seems more appealing.
But as people start going home, the party dies down and more people are encouraged to leave so the tense hosts can clean up.
What is Hills coefficient regarding cooperative enzymes?
MCAT concept check enzyme kinetics page 64 question 1
What are the effects of increasing [S] on enzyme kinetics? Increasing [E]?
Increasing [S] has different effects: when [S] is low, and increase in [S] causes a proportional increase in enzyme activity. At high [S], increasing [S] has no effect on Vmax because Vmax has already been attained.
Increasing [E] will always increase Vmax regardless of starting [E].
MCAT concept check enzyme kinetics page 64 question 2
How are LW-B and M-M graphs similar, how are they different?
MCAT concept check enzyme kinetics page 64 question 3
What does Km represent? What would increase Km significantly?
Km is a measure of an enzymes affinity for its substrate and is defined as the substrate concentration at which an enzyme is functioning at half of its maximal velocity.
As Km increases, an enzymes affinity for its substrate decreases. As Km decreases, an enzymes affinity for its substrate increases.
MCAT concept check enzyme kinetics page 64 question 4
What do the x and y intercept of the LW-B graph represent.
X intercept: -1/Km
Y intercept: 1/Vmax
MCAT concept check enzyme kinetics page 64 question 5
What is enzyme cooperativity?
What is the difference between enzyme activity, enzyme velocity, and enzyme rate?
They are all the same thing.
What three environments influence activity of an enzyme?
T, pH, salinity
How does temperature impact enzyme activity?
Usually double activity per increase in 10°C in temperature up to an optimal T where activity falls off due to denaturation. 37°C, 98.6°F, 310K in humans.
Why are Siamese cats paws, faces, ears, and tails dark colored when the rest of their body is colored white?
How does pH impact enzyme activity? For enzymes in the blood, what is the optimal pH?
Most enzymes depend on pH in order to function properly as it can affect the ionization of the active site as well as denature the protein.
Enzymes in the blood operate best at pH=7.4. pH less than 7.35 is considered acidemia.
What is the optimal pH for enzymes in the blood? Stomach? Duodenum?
Blood: 7.4
Stomach (pepsin): 2
Duodenum (trypsin): 8.5
How does salinity impact enzyme activity?
Increasing levels of salt in vitro (in glass) can disrupt H and ionic bonds, causing a partial change in the confirmation of an enzyme. Can also cause denaturation.
MCAT concept check effects on enzyme activity page 66 question 1
MCAT concept check effects on enzyme activity page 66 question 2
What is feedback regulation? Feed forward regulation? Negative feedback?
Feedback regulation regulates enzymes by products further down a metabolic pathway.
Feed forward regulation: less common. enzymes regulated by intermediates that precede the enzyme in the pathway.
Negative feedback (feedback inhibition): far more common. Helps maintain homeostasis, once we have enough of a given product, it turns off the pathway that creates the product.
Where do we often see negative feedback?
Enzymology and endocrine system. Hormonal feedback loops are also inhibited by negative feedback.
What are the four types of reversible inhibition?
Competitive inhibition: involves occupancy of the active site. Substrates cannot access enzymatic binding sites if there is an inhibitor in the way.
Non-competitive inhibition: binds to an allosteric site instead of the active site, which induces a change in enzyme confirmation.
Mixed inhibition: results when an inhibitor combined to either the enzyme or the enzyme substrate complex, but has different affinity for each.
Uncompetitive inhibition: bind only to the enzyme substrate complex and essentially lock the substrate in the enzyme.
What is competitive inhibition?
Does adding a competitive inhibitor alter the value of Vmax?
Does a competitive inhibitor change Km?
What is the Lineweaver Burk plot of competitive inhibition?
Competitive inhibition simply involves occupancy of the active site. Substrates cannot access and enzymatic binding sites if there is an inhibitor in the way.
Can be overcome by adding more substrate so that the substrate inhibitor ratio is higher.
Competitive inhibitors decrease the value of Km because the substrate concentration has to be higher to reach half the maximum velocity in the presence of the inhibitor.
Adding a competitive inhibitor does not alter the value of Vmax because if enough substrate is added, it will out compete the inhibitor and be able to run the reaction at maximum velocity.
What is non-competitive inhibition?
Can this be overcome by adding more substrate?
Does non-competitive inhibition decrease the value of Vmax?
Does non-competitive inhibition change Km?
What is the Lineweavee Burk plot of non-competitive inhibition?
Non-competitive inhibitors binds to an allosteric site instead of the active site, which induces a change and enzyme conformation.
Because the two molecules do not compete for the same site, inhibition is considered non-competitive and cannot be overcome by adding more substrate. Once the enzyme’s comformation is altered, no amount of extra substrate will be conducive to forming an enzyme substrate complex.
No competitive inhibitors do not change Km because any copies of the enzyme that are still active, maintain the same affinity for their substrate.
Adding a non-competitive inhibitor decreases the measured value of Vmax because there is less enzyme available to react.
What is mixed inhibition?
Mixed inhibition results when an inhibitor combined to either the enzyme or the enzyme substrate complex, but has different affinity for each.
If the inhibitor had the same affinity for both, it would be a non-competitive inhibitor.
Does a mixed inhibitor alter experimental value of Km? Vmax?
Where would the line for inhibitor and no inhibitor converge on a Lw-B plot?
Depends on the preference of the inhibitor for the enzyme versus the enzyme substrate complex.
If the inhibitor preferentially binds to the enzyme, it increases the Km value (lowers affinity).
If the inhibitor preferentially binds to the enzyme substrate complex, it lowers the Km value (increases affinity)
In either case, Vmax is decreased.
The curve for the activity with and without the inhibitor intersect at a point that is not on either axis.
What is uncompetitive inhibition?
What impact do they have on Km, Vmax?
What would a Lw-B plot look like for an uncompetitive inhibitor?
Uncompetitive inhibitors bind only to the enzyme substrate complex and essentially lock the substrate in the enzyme, preventing its release.
Uncompetitive inhibitors lower Km and Vmax.
On a Lw-B plot, the curves for activity with and without an uncompetitive inhibitor are parallel.
Compare reversible inhibitors by binding site, impact on Km, and impact on Vmax.
What is irreversible inhibition?
Irreversible inhibition is when the active site is made unavailable for a prolonged period of time, or the enzyme is permanently altered. This type of inhibition is not easily overcome or reversed.
Aspirin is good example. Acetylsalicylic acid irreversibly modifies cyclooxygenase-1. The enzyme can no longer bind substrate to make its products (prostaglandins) which are involved in modulating pain and inflammatory responses. To make more prostaglandins, new cyclooxygenase-1 we have to be synthesized through transcription and translation.
Irreversible inhibition is a prime drug mechanism, and something you will come across often in medical school.
What are the three kinds of regulated enzymes?
Allosteric. Enzymes that are allosteric have multiple binding sites.
Covalently modded enzymes. Enzymes can be activated or deactivated through covalent modification such as phosphorylation or dephosphorylation.
Zymogens. Enzymes are sometimes secreted as inactive zymogens like trypsinogen.
What is an allosteric enzyme? What is an allosteric activator? Allosteric inhibitor?
What shape will a cooperative steric enzyme have on a Michaelis Menten plot?
Enzymes that are allosteric have multiple binding sites. The active site is present, as well as at least one other site that can regulate the availability of the active site.
Allosteric enzymes alternate between an active and an inactive form. The inactive form cannot carry out enzymatic reaction.
An activator will result in a shift that makes the Active site more available for binding to the substrate.
An inhibitor will make the active site less available.
Michaelis Menten plots of cooperative, allosteric enzyme kinetics often have a sigmoidal curve.
What are covalently modified enzymes? What is glycosylation?
Enzymes are often subject to covalent modification such as being activated or deactivated by phosphorylation or dephosphorylation.
Glycosylation is the covalent attachment of sugar moieties to enzymes. Glycosylation prepare enzyme for transport within the cell, or can modify protein activity.
What are zymogens?
Certain enzymes are particularly dangerous if they are not tightly controlled. These include the digestive enzymes like trypsin, which if released from the pancreas in an uncontrolled manner would digestive the organ itself.
Zymogens are inactive forms of enzymes. They contain a catalytic (active) domain and a regulatory domain. The regulatory domain must be either removed or altered to expose the active site.
Most Zymogens have a suffix -ogen.
MCAT concept check regulation of enzyme activity page 71 question 1
what is feedback inhibition?
Feedback inhibition refers to the product of an enzymatic pathway turning off enzymes further back in the same pathway. This helps maintain homeostasis: as product levels rise, the pathway creating that product is appropriately down regulated.
MCAT concept check regulation of enzyme activity page 71 question 2
Of the four types of reversible inhibitors, which could potentially increase Km?
A competitive inhibitor increases Km because the substrate concentration has to be higher to reach half the maximum velocity in the presence of the inhibitor.
A mixed inhibitor will increase Km only if the inhibitor or preferentially binds to the enzyme over the enzyme substrate complex.
MCAT concept check regulation of enzyme activity page 71 question 3
What is irreversible inhibition?
Irreversible inhibition refers to the prolonged or permanent and activation of an enzyme, such that it cannot be easily re-natured to gain function.
MCAT concept check regulation of enzyme activity page 71 question 4
What are some examples of transient and covalent enzyme modifications?
Example of transient modifications include allosteric activation or inhibition.
Example of covalent modifications include phosphorylation and glycosylation.
MCAT concept check regulation of enzyme activity page 71 question 5
Why are some enzymes released as zymogens?
Zymogens are precursor of active enzymes. It is critical that certain enzymes remain inactive until arriving at their target site.
Digestive enzymes of the pancreas or a great example (trypsinOGEN and chymotrypsinOGEN)
MCAT Mastery Enzymes page 46 question 1
MCAT Mastery Enzymes page 46 question 2
MCAT Mastery Enzymes page 46 question 3
What is an apoenzyme?
An enzyme devoid of its necessary cofactor is called an apoenzyme.
MCAT Mastery Enzymes page 46 question 4
MCAT Mastery Enzymes page 46 question 5
MCAT Mastery Enzymes page 46 question 6
MCAT Mastery Enzymes page 46 question 7
MCAT Mastery Enzymes page 47 question 8
When Km equals the [S], then the enzyme is working at 1/2 Vmax.
MCAT Mastery Enzymes page 47 question 9
MCAT Mastery Enzymes page 47 question 10
This is a good question to reading Lw-B plots.
MCAT Mastery Enzymes page 47 question 11
MCAT Mastery Enzymes page 47 question 12
MCAT Mastery Enzymes page 47 question 13
MCAT Mastery Enzymes page 47 question 14
MCAT Mastery Enzymes page 47 question 15
Relevant equations and a few AA that we need to memorize.
