Worksheetsbiochem exam 2
Total questions: 97
Worksheet time: 3hrs 14mins
Hydrogen bonding helps to stabilize all of the structural levels of protein except for...
Primary structure
Secondary structure
Tertiary structure
Quaternary structure
Quaternary structure
The hydrophobic effect helps to stabilize which of the structural levels of protein below?
Primary structure
Secondary structure
Tertiary structure
Quaternary structure
Tertiary & Quaternary
Which of the following is true for both alpha helices and beta sheets?
Formation of these motifs are enthalpically stabilized by hydrogen bonding
Compared to unfolded polypeptides, their formations are entropically unfavored.
Formation of these motifs are enthalpically stabilized by the hydrophobic effect.
All of above
True or false: Tertiary and quaternary structure are never stabilized by covalent bonds
True
False
Which of the following is the least likely to denature a protein?
Addition of sodium dodecyl sulfate to the protein solution
Boiling the protein solution
Addition of 10 M NaOH to the protein solution
Addition of benzene to the protein solution.
Addition of 1 M NaCl to the protein solution
Which of the following provide no enthalpic contribution to the stability of the 3D structure of a protein?
Hydrophobic effect
Disulfide bonds
London’s forces
Dipole-dipole interactions
Ion-dipole interactions
What concentration of ligand ([L]) is needed for 80. % of the binding sites to bind ligand if the Kd for ligand binding is 500 μM? The protein is a homodimer that exhibits no cooperativity between binding sites.
250 μM
500 μM
2000 μM
200,000 μM
Which secondary structure motifs can be observed in
cytochrome P450? Its protein structure is shown on the right
α-helices
Parallel β-sheets
Antiparallel β-sheets
α-helices and β-sheets
The enzyme cytochrome P450cam contains a heme cofactor (see figure on right question above) and catalyzes oxidation of camphor:
Camphor + NADPH + H+ + O2 -> 5-exo-hydroxycamphor + NADP+ + H2O
Which of the following correctly classifies the NADPH and the heme cofactor?
NADPH is a co-substrate and heme is a prosthetic group
NADPH is a prosthetic group and heme is a metal ion cofactor
NADPH is a co-substrate and heme is a co-substrate
NADPH is a co-substrate and heme is a co-substrate
You are given a protein that is an α2 homodimer (i.e., the subunits that make up the protein are identical). Which SDS-PAGE lane in the figure to the right would be expected for this protein?
A
B
C
D
Which lane in the figure to the right would be expected if SDS were not added to the sample prior to running the experiment? (α2 homodimer (i.e., the subunits that make up the protein are identical))
A
B
C
D
Which of the following occurs when O2 binds to a subunit of hemoglobin?
O2 binding moves the F helix, which weakens ion pairs at the subunit interface and shifts the equilibrium
towards the R state.
O2 binding initiates the Bohr effect, which shift the equilibrium towards the R state.
O2 acts as an H-bond acceptor with the distal histidine, which shifts the equilibrium to the T state.
O2 acts as a base to deprotonate the distal histidine, which shifts the equilibrium to the R state.
The figure below shows linearized Hill plots for four
different ligand binding proteins. Based on the plot,
which protein—A, B, C, or D—shows a Hill lot
consistent with negative cooperativity?
A
B
C
D
On the right is a reaction coordinate diagram for the binding of ligand L to a protein P to form the protein-ligand complex, PL. Based on the reaction coordinate, what is the ΔG for formation of PL?
-5 kJ/mol
-10 kJ/mol
5 kJ/mol
10 kJ/mol
Using the same reaction coordinate diagram that you used in the problem above, what is the activation energy for formation of PL?
-5 kJ/mol
-10 kJ/mol
5 kJ/mol
10 kJ/mol
There is a variant of protein P called protein Q. The ΔG for binding of ligand L by protein Q is 0 kJ/mol. Which of the following statements are true?
The protein Q has a ΔG of 0 kJ/mol and therefore does not bind ligand L.
The Kd for binding of L to protein Q was increased compared to the Kd for binding of L to protein P
The Kd for binding of L to protein Q was decreased compared to the Kd for binding of L to protein P
Neither the forward reaction (association to form the Q-L complex) nor the reverse reaction (dissociation to
form separated L and Q) are thermodynamically favored.
For an irreversible elementary reaction
A + B -> C
What are the units of the rate constant for this reaction?
M-2·s-1
M-1·s-1
s-1
M·s-1
Data for ligand binding curves for a wild-type protein P and a single amino acid mutant of this protein are shown in the table below. What is the effect of the mutation on ligand binding?
The mutation decreases the Kd for ligand binding.
The mutation increases the Kd for ligand binding.
The mutation has no effect on the Kd for ligand binding and only affects the maximum theta value.
A protein P can bind 4 different substrates—T, U, V, or W. The rate constants for the forward (k1) and reverse (k-1) reaction of these equilibria are shown below.
T
U
V
W
The figure on the right displays electrostatic interactions between the α1/β2 subunit interface of hemoglobin. These interactions stabilize the T state of the protein. Which of the following is the most likely effect of removing the Cl - ion from the site?
Increase in the Kd for O2 binding
Decrease in the Kd for O2 binding
No change in the Kd for O2 binding
A protein is ran on size exclusion chromatography and shown to have a molecular weight of 150,000 Da. The same protein is then analyzed by SDS-PAGE with (+BM) and without (-BM) beta-mercaptoethanol. The SDS-PAGE is shown below. The lane labeled "MW" contains a molecular weight marker with the molecular weights labeled on the side. The lane labeled +BM shows the SDS-PAGE for the protein sample containing beta-mercaptoethanol and the lane labeled -BM shows the SDS-PAGE for the protein sample without beta-mercaptoethanol. How many subunits does the protein contain?
(a)
The following sequence is part of a globular protein and form a hydrophobic contact with another part of the protein. Based on the sequence, predict the type of secondary structure.
RFTINLEAHWDLKYQV
Beta-sheet
beta-turn
Alpha-helix
The picture below shows several interactions (shown as dashed lines) that are observed in the crystal structure of a protein. Which of the dashed lines shown in the figure below are likely hydrogen bonds?
D
E
A
B
C
Views from two angles of the same protein are shown below. Estimate the molecular weight of the protein to the nearest 1000 Da.
(a)
A mutation causes all of the glycine residues in the GXY repeats of collagen (See
textbook for more information) to be changed to Phe. Which levels of protein structure
would be most affected by such a mutation?
Primary
Secondary
Tertiary
Quaternary
Protein structure dictates its function
True
False
Primary structure is the amino acid sequence
True
False
A protein consisting of only one peptide chain can exhibit quaternary structure.
True
False
A peptide bond is an amide functional group
True
False
The C and N atoms that compose a peptide bond each have partial sp2 hybridization.
True
False
All of the bonds in the peptide backbone are free to rotate.
True
False
In the Ramachandran Plot shown in Figure 6.9, most of the plot is empty indicating that there are few or no examples of peptides with Phi/Psi angles in those regions. Why are most of the regions empty?
Steric interactions prohibit those Psi/Phi angle pairs.
Resonance prevents those Psi/Phi pairs
Restricted rotation prevents those Psi/Phi pairs
H-bonding is stronger in the allowed regions.
For an alpha helix, how many residues are there per turn?
(a)
An alpha-helix has a dipole moment along the axis of the helix. What type of partial charge is there on the C-terminal end of the helix?
Partial positive charge
Partial negative charge
Adding which of the following to a solution of water-soluble protein might cause the protein to denature?
Sodium dodecyl sulfate (SDS)
Ammonium sulfate
Guanidinium hydrochloride
Acid (e.g., HCl or H2SO4)
A cofactor is part of the polypeptide chain.
True
False
Which of the following are differences between myoglobin and hemoglobin. Only two are correct.
Hemoglobin is the O2 carrier in blood
Myoglobin is the O2 carrier in blood.
Hemoglobin has quaternary structure.
Myoglobin has quaternary structure
A researcher is trying to identify potential ligands for a protein, P. She determines the equilibrium dissociation constants (Kd) for 4 proposed ligands--L, M, N, and O:
Which of the proposed ligands binds the tightest to protein P?
L
M
N
O
For the reaction:
Where the rate constant of the forward reaction is k1 and the rate constant of the reverse reaction is k2, the equilibrium dissociation constant (Kd) is equal to what? Only 3 are correct.
k1/k2
k2/k1
1/Ka
For the reaction...
Where the rate constant of the forward reaction is k1 and the rate constant of the reverse reaction is k2. This reaction will be at equilibrium when
k1 = k2
[A]eq = [B]eq[C]eq
k1[A] = k2[B][C]
Ligand-binding proteins tend to be specific. How does the protein promote ligand specificity?
Controlling the shape of the binding site to match the "shape" of the molecule.
Controlling the shape of the binding site to exclude other molecules.
By charging cover at the door.
The heme in myoglobin is which of the following?
Cofactor
Co-substrate
Co-enzyme
Prosthetic group
Which of the following correctly describes O2 binding to myoglobin?
O2 binding to myoglobin is enthalpically favorable
O2 binding to myoglobin is entropically favorable
O2 binding to myoglobin is enthalpically unfavorable
O2 binding to myoglobin is entropically unfavorable
Which of the following are enthalpic contributions to O2 binding to myoglobin?
O2 binding to iron is endothermic
O2 binding to iron is exothermic
H-bonding of the distal His to O2 is exothermic
H-bonding of the distal His to O2 is endothermic
Using a technique called site-directed mutagenesis, we can change amino acids in a protein from from one type to another. The original protein is typically referred to as the "wild type" or "native" protein. Proteins that are engineered by site-directed mutagenesis to change amino acids are known as "mutants" or "variants". Let us say we engineered a variant myoglobin to replace the distal histidine (the histidine above the O2 binding site) with alanine (H-->A mutation). What would happen to the equilibrium dissociation constant (Kd) of the H-->A mutant compared to the wild-type myoglobin?
The equilibrium dissociation constant (Kd) for O2 binding would increase.
The equilibrium dissociation constant (Kd) for O2 binding would decrease.
The equilibrium dissociation constant (Kd) for O2 binding would remain the same.
What is theta?
The amount of protein-ligand complex
The ratio of protein-ligand complex vs. total protein.
The amount of protein without ligand bound
The ratio of protein-ligand complex vs. free protein
Why does the O2 ligand binding curve approach an asymptote?
Because at high ligand concentrations, the amount of ligand bound is is limited by the amount of protein. The protein gets saturated.
Because ligand-binding curves are log plots.
Because the protein denatures.
What percentage of myoglobin sites have O2 bound when the partial pressure of O2 is 2 torr?
p50 = 2.8 torr
(a)
At what pO2 (in Torr) are 80% of the myoglobin sites occupied with O2?
p50 = 2.8 torr
(a)
What is theta equal to at a ligand pressure of 500 torr if Kd is also 500 torr?
(a)
The ligand binding curves for the binding of three different ligands—L, M, and N—to a protein, P, are shown on the graph below. Based on the binding curves, which ligand—L, M, or N—has the lowest affinity for the protein?
L
M
N
The ligand binding curves for the binding of three different ligands—L, M, and N—to a protein, P, are shown on the graph below. Based on the binding curves, which ligand—L, M, or N—has the lowest affinity for the protein?
L
M
N
What is your estimate for the Kd of ligand L for protein P?
(a)
Why can myoglobin not replace the function of hemoglobin in blood?
Myoglobin has low affinity for O2.
Myoglobin has high affinity for O2.
Myoglobin does not bind O2.
Myoglobin does not exhibit cooperativity necessary to hemoglobin function.
For a Hill plot of myoglobin, what would the term n equal?
(a)
What are the axes for a Hill plot?
Θ vs. initial ligand concentrations
1/Θ vs. initial ligand concentrations.
vs. ligand concentration
vs. log(ligand concentration)
d-2,3-bisphosphoglycerate (BPG) binds to the central cavity of the T-state of hemoglobin (left panel), thereby stabilizing the T state of hemoglobin, which has lower affinity for O2. What type of effector is BPG?
BPG is not an effector
BPG is a positive homotropic effector
BPG is a positive heterotropic effector
BPG is a negative homotropic effector
BPG is a negative heterotropic effector.
The T-state of hemoglobin has a _____(high,low) affinity for O2 while the R state has a _____(high,low) affinity.
(a)
An allosteric effector binds at the same site as the ligand.
True
False
For which model of cooperativity in hemoglobin do all subunits have to be in the R- or the T-state at the same time?
The concerted model
The sequential model
According to the concerted model, all hemoglobin subunits in a solution of 10 μM must be in the R- or T-state at the same time.
True
False
The picture of hemoglobin (the x-ray crystal structure shown in Question 1) was taken at pH 7.2. The pKa of free histidine (i.e., unattached to a protein) is 6.0. Assuming all of the beta2 H146 residues are positively charged at pH 7.2, what is your estimate for the pKa of this residue?
4.0
6.0
7.2
8.0
Binding of O2 to a O2-binding site on hemoglobin promotes conversion between the T- and R-states of hemoglobin. Ion pairs are electrostatic interactions between positively and negatively charged amino acids. For hemoglobin, some of these ion pairs stabilize the T- state of hemoglobin. Based on what you know about the protonation states of amino acids in response to the changes in pH, how will the equilibrium between the T- and R-states of hemoglobin be affected by changes in pH?
An increase in 1 pH unit will further stabilize the T- state, pushing the equilibrium towards the T-state.
An increase in 1 pH unit will destabilize the T-state shifting the equilibrium towards the R-state
A change in pH will have no effect because the ionic charges are permanent and independent of pH
Explain why you do or do not think that the pKa of the beta2-His146 in hemoglobin changes relative to free His?
The negative charge of the nearby D94 increases the pKa of H146
The histidine is incorporated into a peptide bond, making it more basic.
The positive charge on K40 stabilizes the positive charge of H146.
In the table below, the effects on ligand binding affinity of two amino acid mutations of the protein
streptavidin are shown. Explain what effect each mutation had on the affinity of streptavidin for
biotin.
S45A increased the affinity of streptavidin for biotin
D128A increased the affinity of streptavidin for biotin
S45A increased the affinity of streptavidin for biotin
D128A reduced the affinity of streptavidin for biotin
S45A reduced the affinity of streptavidin for biotin
D128A increased the affinity of streptavidin for biotin
S45A reduced the affinity of streptavidin for biotin
D128A reduced the affinity of streptavidin for biotin
An enzyme that converts (S)-lysine to (R)-lysine would be what class of enzyme?
Isomerase
Hydrolase
Ligase
Transferase
Oxidoreductase
Which of the following are properties of enzymes?
Is a catalyst.
Does not get consumed by the reaction
Increase the equilibrium constant for a reaction
Increase the rate of reaction
Made of protein
An enzyme that uses NADH as a co-substrate is most likely what class of enzyme?
Oxidoreductase
Hydrolase
Isomerase
Ligase
Lyase
how many transition states are shown?
(a)
how many intermediates are shown?
(a)
A reaction with ΔG is -10,000 kJ/mol will always proceed to completion in less than 10 s
True
False
How many elementary steps of the reaction are bimolecular (i.e., has a molecularity of 2).
(a)
Concentration of reactants affects which of the following?
Free energy change of a reaction
Rate of a reaction
Q
Keq
Rate constant of a reaction
Temperature affects which of the following?
Free energy change of a reaction.
Rate of a reaction
Rate constant of a reaction
Keq
An enzyme that catalyzes the degradation of tryptophan is likely to catalyze the degradation of other aromatic amino acids.
True
False
If an enzyme can catalyze a transformation of L-isoleucine but not R-isoleucine, this is referred to as
Substrate specificity
Regiospecificity
Product specificity
Which of the following helps the enzyme to confer substrate specificity in enzymes?
Sterics
H-bonds
Electrostatic interactions
Hydrophobic patches
In order to decrease the activation barrier for a reaction, it is most important for an enzyme to complement the
all reaction intermediates.
transition state.
substrate.
product.
What type of enzyme catalyzes the following reaction?
Kinase
Oxidoreductase
Transferase
Ligase
The hypothetical elementary reaction 2A → B + C has a rate constant of 3.0 x 106 M−1 ∙ s−1. What is the reaction velocity (in M/s) when the concentration of A is 15.0 mM?
(a)
An uncatalyzed reac�on has a rate of 4.2 × 10–7 sec–1. When an enzyme is added the rate is 3.2 × 10 4 sec–1
Calculate the rate enhancement caused by the enzyme. Format 0.00E00
(a)
By definition, prosthetic groups are ___ associated with the enzyme polypeptde chain.
tightly
loosely
Even though the proximity effect can be responsible for great portion of the rate
enhancement achieved by enzymes, the most important strategy that enzymes are believed to use rely
on their greater affinity to the reaction ___.
transition state
products
reactants
Which of the following amino acid residues would not provide a side chain for acid-base catalysis at
physiological pH? (Assume pK values of each amino acid are equal to the pK value for the free amino
acid in solution.)
leucine
lysine
aspartic acid
histidine
The covalent catalytic mechanism of an enzyme depends on a single active site Cys whose pK is 8. A
mutation in a nearby residue alters the microenvironment so that this pK increases to 10. Would the
mutation cause the reaction rate to...
increase
decrease
Classify the enzymes catalyzing the following reactions according to the 6 classes
Isomerase
Lyase
Oxidoreductase
Ligase
Classify the enzymes catalyzing the following reactions according to the 6 classes
Isomerase
Lyase
Oxidoreductase
Ligase
Classify the enzymes catalyzing the following reactions according to the 6 classes
Isomerase
Lyase
Oxidoreductase
Ligase
Classify the enzymes catalyzing the following reactions according to the 6 classes
Isomerase
Lyase
Oxidoreductase
Ligase
What variables need to be controlled when monitoring reaction progress?
Select all that apply.
Temperature
pH
Concentration of reactants
How would you ensure the pH does not change over the course of an enzymatic reaction?
Only study reaction that do not use or generate protons.
Use a buffer
Use only water as the solvent.
Add acid to the reaction mixture
Add base to the reaction mixture
Based on what you know about how temperature can influence a reaction, why must temperature be controlled over the course of an enzyme experiment?
Temperature influences the 3D structure of an enzyme.
Temperature influences the rate constant of a reaction.
Temperature influences the rate of the reaction.
Temperature influences the equilibrium of a reaction.
Based on what you know about how reactant concentration can influence a reaction, why must concentration of reactants be controlled in an enzymatic experiment?
Reactant concentration influences the 3D structure of an enzyme.
Reactant concentration influences the equilibrium of a reaction.
Reactant concentration influences the rate constant of a reaction.
Reactant concentration influences the rate of the reaction.
What are the x- and y-axes of a linearized plot of a first-order reaction?
x-axis: ln(time); y-axis: ln([A]/[A0])
x-axis: time; y-axis: [A]/[A0]
x-axis: time; y-axis: ln([A]/[A0])
x-axis: time; y-axis: 1/[A]
What are the x- and y-axes of a linearized plot of a second-order reaction where both reactants are the sample compound? In other words, the reaction is
2A --> B
x-axis: time; y-axis: ln([A]/[A0])
x-axis: ln(time); y-axis: ln([A]/[A0])
x-axis: time; y-axis: [A]/[A0]
x-axis: time; y-axis: 1/[A]
What are the units of the rate constant that you found in (First order rxn)?
M-1s-1
s-1
Which of the following are assumption made when deriving the Michaelis-Menten equation?
The steady-state approximation.
That any change in substrate concentration is negligible and the substrate concentration is effectively constant.
There are two intermediates.
Substrate binding to enzyme is irreversible.
