384 Exam 2
First view the ribbon structure of a subunit of the human aquaporin. Next, change the display of the amino acids in the "a (amino)" polypeptide at the constriction point to ball-and-stick or space-fill. The constriction point is represented as a blue space-fill model in the default view. To change the display, first select "a (amino)" on the bottom of the player screen by clicking the green-outlined box next to it. From the dropdown menu, click the icon for space-fill or ball-and-stick on the Display line. To remove the green x's, click the icon for "Clear Selection" on the Misc line in the dropdown menu. Identify the amino acids at the constriction point. The constriction point within the water channel of aquaporin proteins is formed by two short α helices, each of which contains a conserved ______________ residue that interacts with translocating water molecules.
asparagine
Although most protein sequencing is now done by mass spectrometry (MS), Edman degradation (ED) is useful if large amounts of a protein can be obtained even if from an uncharacterized species. Answer the True/False questions regarding methods used to sequence a polypeptide fragment and choose the set of all correct answers. 1. True / False - Protein sequencing by ED does not require purified protein but sequencing by MS does. 2. True / False - The N-terminal residue removed by acid treatment in ED is always a methionine. 3. True / False - Trypsin and chymotrypsin treatment often provides identical peptide fragments for ED. 4. True / False - Genomic sequences are needed to predict protein sequences by the MS method. 5. True / False - Trypsin treatment provides useful sequence information when analyzing data from MS.
(1) False (2) False (3) False (4) True (5) True
Consider the reaction above. For the uncatalyzed reaction, kF(uncatalyzed) = 5 X 10-4 s-1 and Keq is 2.5 X 102. For the same reaction in the presence of an enzyme, kF(catalyzed) is 5 X 105 s-1. B.) What is kR(catalyzed)? (IMAGE) A.) 2X103 s-1 B.) 5X105 s-1 C.) 5X10-13 s-1 D.) Incorrect Response E.) 5X104 s-1
A.) 2X103 s-1
Which amino acid acts as a general acid and a general base in the mechanism of chymotrypsin? A.) His57 B.) Gly193 C.) Ser195 D.) Asp102
A.) His57
A mutation in the mc074-1.jpg subunit of hemoglobin is discovered that reduces the affinity of 2,3-BPG binding. Which of the following mutations is most likely to have this consequence? A.) Lys82 --> Asp82 B.) Glu6 --> Val6 C.) Val1 --> Ile1 D.) Asp99 --> Glu99
A.) Lys82 -->Asp82
First, view just the heme cofactor of myoglobin and locate the four amine nitrogen atoms that are in close proximity to the iron ion (bright blue color in the center). Next, click the dropdown menu in the upper right corner and click the "Measure Distance" icon. Now use the angstrom measurement tool to determine the approximate distance between any one of the four amine nitrogen atoms and the iron ion. This distance is approximately ______ angstrom(s).
2 angstrom(s)
Place the following HMG-CoA reductase steps in the correct order: A. Reduction of aldehyde B. Breakdown of hemithioacetal C. Reduction of thioester D. Cofactor exchange 1.) A, D, C, B 2.) C, B, D, A 3.) A, B, D, C 4.) C, D, B, A
4.) C, D, B, A
Calculate the purification of the target protein when there is a 30% decrease in activity and a 55% decrease in total protein after centrifugation. A.) 1.6-fold B.) 0.55-fold C.) 0.64-fold D.) 1.8-fold
A.) 1.6-fold
Which resin should you subject your mixture to get the best separation? Choose the resin with the correct attribute of the resin and protein that will separate the proteins. A.) CMC cellulose, because the pH is above two of the contaminating proteins and less than YPOI and contaminant protein 3. These two proteins can then be separated by ionic conditions (salt gradient). B.) DEAE cellulose, because the proteins will be charged and can be separated by ion exchange chromatography C.) gel filtration, because the proteins are all within the exclusion range of this resin D.) CMC cellulose, because all four proteins will be positively charged and can be separated by ionic conditions during elution
A. CMC cellulose, because the pH is above two of the contaminating proteins and less than YPOI and contaminant protein 3. These two proteins can then be separated by ionic conditions (salt gradient).
A particular genomic caretaker protein, Protein X, has an affinity for both Ligand Y and Ligand Z. When you have 0.23 microM of Protein X in a solution and mix it with 0.11 microM of Ligand Y, the resulting solution contains 0.20 microM of free Protein X, 0.09 microM of free Ligand Y, and 0.02 microM of the protein-ligand complex, after equilibrium has been reached. However, when you have 0.23 microM of Protein X in a solution and mix it with 0.11 microM of Ligand Z, the resulting solution contains 0.14 microM of free Protein X, 0.02 microM of free Ligand Z, and 0.09 microM of the protein-ligand complex, after equilibrium has been reached. Which ligand has a greater affinity for Protein X? A. Ligand Z B. Ligand Y C. It is not possible to determine with the given information. D. Ligands Y and Z have equal affinity for Protein X.
A. Ligand Z
Which of the following statements are true about the poison ouabain? A. Ouabain inhibits the Na+-K+ ATPase protein, and this particular inhibition stops muscles such as the heart and lungs from relaxing after contracting. B. Ouabain inhibits the Na+-K+ ATPase membrane transport protein, which is used to passively transport sodium and potassium ions through the cell membrane. C. Ouabain is a poison found in the seeds of the climbing oleander plant, and is used by African tribesmen in poison arrows. However, in small doses, it can be used to treat patients with heart problems. D. Ouabain is a small molecule that contains 35 carbon atoms and multiple hydroxyl groups decorating a fused ring system. (TWO ANSWERS)
A. Ouabain inhibits the Na+-K+ ATPase protein, and this particular inhibition stops muscles such as the heart and lungs from relaxing after contracting. C. Ouabain is a poison found in the seeds of the climbing oleander plant, and is used by African tribesmen in poison arrows. However, in small doses, it can be used to treat patients with heart problems.
Within this molecular structure, the polyatomic ion sulfate (SO42-) is observed. Which of the following best describes the interaction between myoglobin and this polyatomic ion? Choose one: A. Sulfate was used during precipitation of myoglobin and is seen in this molecular structure interacting with amino acids. B. Sulfate was used during precipitation of myoglobin and is seen in this molecular structure interacting with the heme cofactor. C. Sulfate is part of the normal cellular physiology of myoglobin and is seen in this molecular structure interacting with the heme cofactor. D. Sulfate is part of the normal cellular physiology of myoglobin and is seen in this molecular structure interacting with amino acids.
A. Sulfate was used during precipitation of myoglobin and is seen in this molecular structure interacting with amino acids.
Looking at the figure below, to which of the following does an enzyme bind the strongest? (IMAGE) Choose one: A. Transition state B. ES C. EP D. S
A. Transition state
Which of the following is/are true regarding initial and maximal velocity of enzyme-mediated reactions? Choose one or more: (3 ANSWERS) A. When an enzyme behaves with Michaelis-Menten kinetics, the initial velocity of that enzyme reaction changes with substrate concentration, but the maximal velocity of that particular enzyme remains the same at a constant enzyme concentration and identical reaction conditions. B. At very low substrate concentrations, the initial velocity of an enzyme reaction will be lower than the maximal velocity of the enzyme reaction. C. Initial velocity is maintained until maximal velocity is reached. D. The initial velocity is determined by the slope of the line at the beginning of a reaction when one plots the experimental results of reaction time versus product formed. Maximal velocity is then determined by when the initial velocity no longer significantly changes with increasing substrate concentration.
A. When an enzyme behaves with Michaelis-Menten kinetics, the initial velocity of that enzyme reaction changes with substrate concentration, but the maximal velocity of that particular enzyme remains the same at a constant enzyme concentration and identical reaction conditions. B. At very low substrate concentrations, the initial velocity of an enzyme reaction will be lower than the maximal velocity of the enzyme reaction. D. The initial velocity is determined by the slope of the line at the beginning of a reaction when one plots the experimental results of reaction time versus product formed. Maximal velocity is then determined by when the initial velocity no longer significantly changes with increasing substrate concentration.
Which of the following is not a property or characteristic of both hemoglobin and myoglobin? A. a protein composed of four polypeptide chains B. a protein that contains iron as a metal cofactor C. a protein that contains heme as a prosthetic group D. a protein that binds O2
A. a protein composed of four polypeptide chains
In what direction is water expected to move through this protein? A. from the cytoplasm to the extracellular space B. from the extracellular space to the cytoplasm C. from the extracellular space to the interior of the membrane D. from the cytoplasm to the interior of the membrane (TWO ANSWERS)
A. from the cytoplasm to the extracellular space B. from the extracellular space to the cytoplasm
Shown here is the potassium channel. Consider its molecular structure and manipulate the molecule according to the instructions below to answer the following questions. Display or remove from display the four subunits of the potassium channel by clicking the black-outlined boxes next to where they are listed along the bottom of the player. Study the lengths of the subunits' longitudinal axes in comparison to one another. How many of the four subunits of the potassium channel contain membrane-spanning helices? (Note that the initial view contains the ribbon structure of all four subunits visible.) Choose one: A. one B. two C. three D. four
A. one B. two C. three D. four
Experiments are performed to determine the initial reaction velocity of an enzyme-catalyzed reaction. What is NOT held constant so that the initial velocities can be used to plot the Michaelis-Menten graph? Choose one: A. substrate concentration B. temperature C. enzyme concentration D. volume of reaction
A. substrate concentration
An enzyme has a single active site at which it can bind and hydrolyze either X or Y; however, the enzyme cannot bind X and Y at the same time. Answer the following questions regarding the Km and vmax of this enzyme. (Choose a total of three answers, one each for questions A, B, anc C) A) Will the apparent Km for X be affected if Y is present in the reaction mixture? B) Will vmax for X be affected if Y is present in the reaction mixture? C) Is it possible for vmax and vmax/Km to show a different dependence on pH? (3 CORRECT ANSWERS) A.) NO C.) YES B.) YES A.) YES B.) NO C.) NO
A.) NO B.) YES C.) NO
The figure below shows the coordination of heme, O2, and two critical histidine residues in globin proteins. Which of the following steps happens first in the oxygen binding process? (IMAGE) A.) O2 binds to the iron of heme. B.) The proximal histidine moves toward heme. C.) Iron moves into the plane of heme. D.) The F helix moves toward heme.
A.) O2 binds to the iron of heme.
A mixture of enzyme and inhibitor is run through a size-exclusion chromatography column. The activity of the enzyme is assessed before and after the chromatography. The enzyme has more activity after the chromatography step. Which of the following is true? A.) The inhibitor is a reversible inhibitor. B.) The enzyme was denatured during chromatography. C.) The enzyme was degraded by the column. D.) The inhibitor is an irreversible inhibitor. E.) The enzyme was not eluted fully from the column.
A.) The inhibitor is a reversible inhibitor.
Which of the following would shift the oxygen binding curve for hemoglobin to the left? A.) a higher pH B.) a lower concentration of CO2 C.) an increase in the number of hemoglobin molecules in the T state D.) a higher concentration of 2,3-BPG
A.) a higher pH
Describe one advantage and one disadvantage of trying to solve molecular protein structures by 1) X-ray crystallography and by 2) Nuclear Magnetic Resonance (NMR) spectroscopy.
An advantage of X-ray crystallography is that there is no theoretical size limitation for the proteins being studied. One disadvantage of X-ray crystallography is that protein crystals can be hard to obtain due to how the conditions that may facilitate the crystallization of a protein may be unknown. An advantage of Nuclear Magnetic Resonance (NMR) spectroscopy is that conformational changes or dynamic fluctuations can be observed within proteins, but NMR also has the disadvantage of requiring high protein concentrations of about 0.1-0.5 mM, which may not be possible to have for all proteins.
Which of the following reactions directly alters DNA and affects gene expression? A. Reversible covalent modification involving phosphorylation B. Reversible covalent modification involving methylation C. Coenzyme-dependent redox reactions D. Metabolite transformation reactions
B. Reversible covalent modification involving methylation
here are nine lysine residues within pea gylcine carboxylase. Why would a specific lysine attachment site for lipoamide be conserved among orthologous decarboxylase proteins? Choose one: A. Allosteric regulators prevent lipoamide from attaching at the other lysine residues. B. The lipoamide needs to be precisely located near the enzyme active site. C. Glycine decarboxylase is not subject to selective pressure and is unlikely to undergo mutation. D. Lysine residues in alpha helices have side chains that are sequestered from being able to bind cofactors.
B. The lipoamide needs to be precisely located near the enzyme active site.
Generally, the chemistry of Fmoc blocking is straightforward for most amino acids during solid state peptide synthesis. There is one amino acid, however, that presents a problem for Fmoc blocking during solid state peptide synthesis. That amino acid is A. arginine B. lysine C. glycine D. glutamate
B. lysine
Leghemoglobin is an oxygen-binding protein in root nodules that contain bacteria that fix atmospheric nitrogen. Which ONE of the following is TRUE if leghemoglobin is more like myoglobin than hemoglobin? A.) O2 binding changes the heme configuration from T to R. B.) The O2 binding curve is hyperbolic C.) The O2 binding curve is sigmoidal D.) There is cooperativity in oxygen binding E.) There are four oxygen binding sites
B.) The O2 binding curve is hyperbolic
The size of proteins studied by NMR is limited because A.) the measurements of the distances between atoms are only approximate. B.) large molecules reorient slowly in solution. C.) it is hard to determine the phases of diffracted X-rays. D.) a small sample size is needed.
B.) large molecules reorient slowly in solution.
Though oxygen only binds to a particular place in the hemoglobin molecule, the oxy- and deoxy- forms of hemoglobin differ in overall protein conformation. An example of a structural change that does NOT play a role in translating local binding of oxygen to global protein conformational change would be the different ______ in the presence and absence of oxygen binding. A.) size of the heme iron B.) location of the distal histidine C.) orientation of the F helix D.) location of the proximal histidine
B.) location of the distal histidine
The initial velocity of an enzyme-catalyzed reaction is followed at various substrate concentrations. At very high substrate concentrations it is observed that the initial velocity no longer increases as more substrate is added. The velocity under these conditions is known as A.) the ultimate velocity. B.) the maximum velocity. C.) optimal velocity. D.) v[S].
B.) the maximum velocity
The contractile unit of skeletal muscle is the sarcomere, which is flanked by two Z disks. Sort the proteins listed based on their locations. Between Z disks and Within Z disks Troponin -Tropomyosin -Actin -Myosin Desmin -Vimentin -α-actinin
Between Z disks: -Troponin -Tropomyosin -Actin -Myosin Within Z disks: -Desmin -Vimentin -α-actinin
Now answer this question: Protein secondary structure is important to the function of proteins and consists of three main types: alpha helix, beta strand, and beta turn. What type of protein secondary structure is highlighted? Choose one: A. Beta strand B. Beta turn C. Alpha helix
C. Alpha helix
Which component of a tandem mass spectrometer determines the mass of subfragments? A. Ionization chamber B. Detector C. Mass spectrometer 2 D. Mass spectrometer 1 E. Collision chamber
C. Mass spectrometer 2
The first step of protein mass spectrometry is to get the protein (usually peptide fragments) into a gas phase as an ion. Which of the following describes matrix-assisted laser desorption/ionization (MALDI) ionization? A. Trypsinized fragments can be exposed directly to the first chamber of a tandem mass spectrometry chamber, where collisions with gas particles cause peptide fragmentation. B. Liquid chromatography is used for elution into a focused laser beam to fragment each eluted protein. C. Protein fragments are embedded in a solid mixture that absorbs light, and then a laser flashes on this mixture, leaving fragmented and ionized peptides in the gas phase. D. Small peptide fragments are released from a small metallic capillary under high voltage that removes the solvent and ionizes the peptides into the gas phase.
C. Protein fragments are embedded in a solid mixture that absorbs light, and then a laser flashes on this mixture, leaving fragmented and ionized peptides in the gas phase.
Using immunoprecipitation, you can isolate a protein (protein X) you think is involved in chronic myelogenous leukemia (CML), which is caused, in part, by a hyperactive tyrosine protein kinase called ABL. You think one of the targets of ABL is the protein X that you can purify. In comparing tissues with and without the disease, you subject the samples to isoelectic focusing. Understanding how IEF works, how would you expect the samples with CML to migrate compared to the wild-type, non-diseased sample protein? Use the figure to help you consider your response. A. The non-phosphorylated proteins from the non-diseased samples will move further to the anode without the bulky phosphate group attached to a tyrosine. B. The phosphorylated protein will be more attracted to the negative charge on the anode. C. The phosphorylated protein will have a lower isoelectric point and thus migrate further toward the anode. D. Non-diseased proteins will migrate with a lower pI than those from diseased tissues and thus the samples from the non-diseased samples will migrate more to the anode than diseased protein.
C. The phosphorylated protein will have a lower isoelectric point and thus migrate further toward the anode.
Which of the following is not an assumption that is made when applying Michaelis-Menten kinetics to an enzyme? Choose one: A. The concentration of ES is relatively constant after the initial reaction time. B. The product release is a rapid step in the process. C. The rate constant of E + P re-associating to form the ES complex must be considered. D. The reaction must be considered early, before any appreciable amount of product has been generated.
C. The rate constant of E + P re-associating to form the ES complex must be considered.
Which of the following is correct about turnover number (kcat) and the specificity constant for an enzyme? Choose one: A. kcat = vmax/[ES] B. The specificity constant is defined as (kcat)(Km). C. The specificity constant is defined as kcat/Km. D. The kcat reveals how well an enzyme works.
C. The specificity constant is defined as kcat/Km
Trypsin, chymotrypsin, and elastase are all serine proteases that cleave after different amino acids. What is responsible for the substrate specificity? Choose one: A. Different amino acids involved in the catalytic triad B. Each protease is made in a different cell type. C. The substrate binding pockets accommodate different amino acids. D. Different catalytic mechanisms
C. The substrate binding pockets accommodate different amino acids.
Which of the following statements are true regarding hemoglobin? Choose one or more: A. The T state of hemoglobin has a higher affinity for oxygen than the R state. B. When hemoglobin shifts from the T-state conformation to the R-state conformation, many covalent interactions are broken and reformed. C. When hemoglobin shifts from the T-state conformation to the R-state conformation, many noncovalent interactions are broken and reformed. D. The R state of hemoglobin has a higher affinity for oxygen than the T state.
C. When hemoglobin shifts from the T-state conformation to the R-state conformation, many noncovalent interactions are broken and reformed. D. The R state of hemoglobin has a higher affinity for oxygen than the T state.
Identify the location within myoglobin where O2 interacts. Which best describes where and how O2 interacts in myoglobin? Choose one: A. with an amino acid sequence within a beta sheet of the protein B. on both sides of the heme cofactor and associated with amine nitrogen atoms C. on one side of the heme cofactor and associated with the iron ion D. with the negatively charged carboxy terminus of the polypeptide
C. on one side of the heme cofactor and associated with the iron ion
Watch the animation and then answer the question. The mass-to-charge ratios of denatured proteins are equivalent for different mass proteins. However, the cross-linked nature of the acrylamide media can limit migration through the polymer matrix. Gels with less cross-linked acrylamide (low % SDS gels) will do which of the following? A. separate the smaller proteins based on the percentage gel at the expense of larger proteins B. separate proteins as in size-exclusion chromatography, with the smaller proteins migrating most slowly and the larger proteins migrating farther through the gel C. separate larger proteins at the expense of smaller proteins, which will not resolve well D. allow the negative charge contributed by the smaller SDS molecules to more easily migrate through a low-percent acrylamide because of the increase in negative charge
C. separate larger proteins at the expense of smaller proteins, which will not resolve well
Select the one TRUE statement regarding how catalysts increase reaction rates A.) Catalysts lower the ground state energy of the product, thereby lowering deltaG of the reaction. B.) Catalysts raise the ground state energy of both the product and reactant, thereby lowering the activation energy C.) Catalysts lower the amount of energy required to reach the transition state. D.) Catalysts lower the ground state energy of both the product and reactant.
C.) Catalysts lower the amount of energy required to reach the transition state.
One of the most difficult steps in X-ray crystallography is A.) obtaining a large enough sample for analysis. B.) mounting the crystal on the x-ray diffractor. C.) determining the phases of diffracted X-rays. D.) dissolving the protein in the appropriate solvent. E.) using the correct radio frequency pulses to perturb the nuclear spin
C.) determining the phases of diffracted X-rays.
In the question above, what is the Kd for Ligand Z and Protein X? Choose one: A. .0511 B. 32.14 C. 19.56 D. 0.031
D. 0.031
Which of the following sample preparation steps is used for both native PAGE and SDS-PAGE? A. Heating sample in boiling water bath B. Addition of β-mercaptoethanol C. Addition of sodium dodecyl sulfate D. Addition of glycerol to the sample loading buffer
D. Addition of glycerol to the sample loading buffer
Which of these statements about transport across cell membranes is false? A. Passive transport across cell membranes occurs through membrane proteins, in the same direction as the concentration gradient. B. Small, nonpolar molecules can often diffuse across a cell membrane, traveling from an area of high concentration of solute to an area of low concentration of solute. C. Active transport proteins translocate biomolecules across the cell membrane, against the concentration gradient. D. Both active and passive transport require the addition of external energy, usually in the form of ATP.
D. Both active and passive transport require the addition of external energy, usually in the form of ATP.
On the 2-D gel shown in the figure below, where would a protein with a high pI and a high mass be found? A. B B. D C. A D. C
D. C
Which of the following is accurate in terms of the relationship between the velocity of a reaction and the rate constant (k) of a reaction? Choose one: A. For both first-order and second-order reactions, the concentration of substrate is equal to the product of the velocity of the reaction and the rate constant of the reaction. B. In a second-order reaction, the rate constant is equal to the velocity of the reaction multiplied by the concentration of both substrates. C. For a first-order reaction, the rate constant of a reaction is equal to the product of the substrate concentration and the velocity of the reaction. D. In a first-order reaction, the rate constant of a reaction is equal to the velocity of the reaction divided by the concentration of substrate.
D. In a first-order reaction, the rate constant of a reaction is equal to the velocity of the reaction divided by the concentration of substrate.
The lysine residue that is integral to the reaction mechanism of aldolase is positioned between a leucine residue and a proline residue. Which of the following is true about this lysine? A. It is one of the residues connecting one α helix to another α helix. B. It is part of a β sheet. C. It is part of an α helix. D. It is one of the residues connecting a β sheet to an α helix. E. It is one of the residues connecting one β sheet to another β sheet.
D. It is one of the residues connecting a β sheet to an α helix.
Lipoamide is a covalently attached coenzyme that plays a key role in several decarboxylase reactions, including glycine decarboxylase, and is attached to a lysine residue in the enzyme. Review the molecule again. Manipulate it in order to identify the specific lysine residue that serves as the attachment site for lipoamide in the enzyme glycine decarboxylase. You may want to view the hint if you need help. Choose one: A. Lysine 118 B. Lysine 17 C. Lysine 8 D. Lysine 63
D. Lysine 63
One step in the reaction mechanism of aldolase is represented in this molecular structure. Which of the following best describes the stage of the aldolase mechanism that is captured here? You may need to rotate the ball-and-stick or the space-filling model of the atomic representation so that you can observe the amino acid side chain. The ribbon structure will provide the least amount of help. Additionally, be sure to view the amino acid sequence and observe that the side chain of interest is flanked by a leucine residue and a proline residue. Choose one: A. The dihydroxyacetone phosphate is noncovalently associated to a lysine side chain. B. The glyceraldehyde-3-phosphate is covalently bound to a lysine side chain. C. The glyceraldehyde-3-phosphate is noncovalently associated to a lysine side chain. D. The dihydroxyacetone phosphate is covalently bound to a lysine side chain.
D. The dihydroxyacetone phosphate is covalently bound to a lysine side chain.
Congratulations, you have purified your protein of interest (YPOI). You and your research partner have subjected your protein to polyacrylamide electrophoresis. You treated your protein with SDS and a reducing agent (beta-mercapto ethanol) and boiled it. Your partner ran a native gel without the SDS and reducing agent treatment. You both observe one band on each gel, but your protein (denaturing SDS PAGE) migrated as a 23,000-dalton protein, while your research partner's gel showed a band at 92,000 daltons. After some discussion, you feel that your protein is a tetramer and that the SDS and reducing agent have: Choose one: A. degraded (hydrolyzed) the protein to smaller peptides. B. folded the protein into a smaller, more compact protein, allowing it to funnel through the acrylamide more easily. C. altered the ability of the protein to migrate through acrylamide. D. denatured the protein, which is made of equally sized monomers.
D. denatured the protein, which is made of equally sized monomers.
sickle cell anemia Choose one: A. results from polymerization of α2βSβ or α2βS2. B. results from polymerization of oxyhemoglobin HbS. C. results from a mutation that puts a charged amino acid on the β subunit protein surface. D. is thought to provide heterozygous individuals protections from malaria.
D. is thought to provide heterozygous individuals protections from malaria.
After centrifugation, there is a 10% decrease in activity and a 75% decrease in total protein. What is purification of the target protein? A.) None of these answers are correct. B.) 7.5-fold C.) 1.3-fold D.) 3.6-fold E.) 0.28-fold
D.) 3.6-fold
The graph above shows three potential oxygen saturation curves for hemoglobin. If the tetrameric hemoglobin is dissociated into its subunit, the saturation curve would change as represented by a shift from... (choose ONE answer) A.) C to B B.) B to C C.) A to C D.) B to A E.) A to B
D.) B to A
In one type of hemoglobin mutant the amino acid change eliminates a hydrogen bond that normally stabilizes the T state. Would you expect this mutant hemoglobin to have a higher or lower O2 affinity compared to the normal protein and why? A.) Lower O2 affinity because the R-T equilibrium would be mostly shifted to the R state. B.) Higher O2 affinity because the R-T equilibrium would be mostly shifted to the T state. C.) Lower O2 affinity because the R-T equilibrium would be mostly shifted to the T state. D.) Higher O2 affinity because the R-T equilibrium would be mostly shifted to the R state
D.) Higher O2 affinity because the R-T equilibrium would be mostly shifted to the R state
Which one of the findings below would provide support for the transition state theory of enzyme catalysis? A.) The activation energy of an enzyme catalyzed reaction is increased, indicating tight binding of reaction intermediates. B.) Products of the reaction slowly dissociate from the enzyme C.) Multiple substrates bind in an ordered fashion (i.e. substrate A binds first, then B, then C) to active sites. D.) Transition state analogs bind tightly to enzyme active sites
D.) Transition state analogs bind tightly to enzyme active sites
Which answer correctly pairs the enzyme class with the type of reaction catalyzed? A.) transferase; transfer of H or O atoms B.) lyase; formation of two products by hydrolyzing a substrate C.) oxidoreductase; transfer of groups within molecules D.) isomerase; intramolecular rearrangements
D.) isomerase; intramolecular rearrangements
What is the rate enhancement as a result of the presence of an enzyme if the uncatalyzed rate of the reaction is 1.2 mc005-1.jpg 102 mmol/sec and the catalyzed rate is 2.4 mc005-2.jpg 104 mmol/sec? A.) 2.88x10^6 B.) 2 C.) 0.005 D.) 20 E.) 200
E.) 200
If the histidine on the F helix (His F8) that coordinates the O2 that binds to the heme iron in hemoglobin is mutated to an alanine, what effect would be most likely? Choose the ONE most correct answer. A.) Oxygen binding affinity would increase, since the alanine residue would form strong interaction with the oxygen molecule. B.) None of these answers are correct. C.) Oxygen binding would cause hemoglobin polymers to form, due to the introduction of a hydrophobic patch by the addition of this alanine. D.) Oxygen binding would take place on the other face of the heme molecule, where the "proximal" histidine would be able to interact with the oxygen. E.) Oxygen binding would not cause the movement of the F helix, since the alanine at F8 cannot coordinate oxygen.
E.) Oxygen binding would not cause the movement of the F helix, since the alanine at F8 cannot coordinate oxygen.
Oxygen binding is monitored for a solution of hemoglobin. During the experiment, the curve changes from sigmoidal to hyperbolic. Which of the following may be the reason for the change? A.) A negative allosteric effector was added. B.) None of these answers are correct. C.) A positive allosteric effector was added. D.) The protein denatured. E.) The protein dissociated into individual subunits.
E.) The protein dissociated into individual subunits.
There are two main methods to get proteins into a gas phase for mass spectroscopy: electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI). Place the appropriate description of each ionization method in the appropriate space.
ESI: -Degrades and charges the peptide with high voltage -Sample becomes ionized due to high voltage. -Ionization evaporates solvent, leaving peptide in gas phase to travel through the separating mass spectrometer. Maldi: -Laser will be absorbed by the peptide containing material releasing the peptide into the gas phase. -Proteins or peptides are mixed in a solid matrix rather than a solution. -Peptides become fragmented and charged due to laser exposure Both ESI and Maldi: -Uses mass-to-charge (m/z) ratio to determine molecular mass -Can be used for proteins or peptides that have been treated with proteases like trypsin -Is based on the mass and acceleration though a chamber ending in a detector
2,3-BPG is a negative allosteric regulator of hemoglobin (Hb) oxygen binding. Explain how elevated levels of 2,3-BPG in red blood cells at high altitude (14,800 ft above sea level) lead to more efficient delivery of O2 to the tissues considering that less O2 binds to Hb in the lungs under these conditions.
Elevated levels of 2,3-BPG in red blood cells at high altitude lead to a more efficient delivery of O2 to the tissues by increasing the amount of O2 deloaded due to a shift in the equilibrium to the T state. Although there is less oxygen saturation, the increase in O2 deloading offsets this, making a higher delivery of O2 to the tissues.
What kinetic parameter reveals the most information about how well an enzyme works? Describe this kinetic parameter in terms of the turnover number and the Michaelis constant and provide an explanation for why triosephosphate isomerase is considered to be 100,000 better than pepsin when this kinetic parameter is used to compare these two enzymes.
Enzyme kinetics is the quantitative analysis of reaction rate data obtained within purified enzymes under lab conditions. The turnover number provides maximum catalytic rate information but does not tell the efficiency of the enzyme meaning that two enzymes will have different reactions but with the same kcat value. Triose phosphate isomerase is 100,000 better than pepsin because the turnover rate is much higher; 4.3 x 103 compared to pepsin which has a 5.0 x 10-1 turnover rate and lower Km.
Three critical features of enzyme structure and function contribute to the overall efficiency of enzymes as biological catalysts and to their central role in biochemical processes. Articulate these three key features and describe a specific enzyme example that illustrates each one.
Enzymes bind substrates with high affinity and specificity. They have an active site that acts as a pocket where the substrate can bind. This active site provides the chemical environment necessary for the catalytic reaction to occur. An example of this is the enzyme phosphoglycerate kinase which contains several key amino acids (arginine) at the binding site which help to orient the substrate (3-phosphoglycerate) through electrostatic interactions. Substrate binding to the active site induces structural changes in the enzyme which result in a large number of weak interactions between the substrate and enzyme. An example of this is the glycolytic enzyme hexokinase which undergoes a large conformation change upon binding of the glucose substrate. This conformational change helps to exclude water from the active site and affects the energy of reaction catalysis. Enzyme activity is highly regulated in cells. Enzyme regulation is required to provide the proper response to environmental stimuli and balance anabolic and catabolic pathways accordingly. An example of this is glycogen phosphorylase which is regulated by regulatory molecules. The catalytic efficiency of this enzyme is increased by the phosphorylation of a serine residue and by the non-covalent binding of adenosine monophosphate.
In addition to O2, another diatomic molecule that can bind to the iron atom in hemoglobin is CO. Explain in biochemical terms why high levels of CO gas are lethal and cite a specific example of accidental death by CO poisoning (where/when/how).
High levels of CO gas are lethal because CO has an extremely high affinity for the heme Fe2+, thereby blocking oxygen delivery to the tissues. One example of accidental death by CO poisoning can be found in Des Moines, Iowa. On May 8, 2020, the man died from carbon monoxide poisoning after he worked on a furnace in a garage.
A genetic mutation that causes which of the following substitutions would be the least likely to destroy the catalytic mechanism that relies upon the lysine residue discussed in Parts 1 and 2 regarding the aldolase enzyme? K to E K to R K to Y K to D K to G
K to R
For each of the five major protein types in eukaryotic cells 1) provide a specific example of a protein in each class, 2) describe its primary function, and 3) identify cellular locations where you would find this specific protein (nucleus, cytosol, membranes).
Metabolic Enzymes: An example of a metabolic enzyme is malate dehydrogenase, which oxidizes malate to create oxaloacetate. The protein can be found in the mitochondrial matrix or the cytoplasm, depending on its isoform. Structural Proteins: An example of a structural protein is collagen, which strengthens the tendons, cartilage, bones, and teeth. Collagen can be found in the extracellular matrix in connective tissue. Transport Proteins: The sodium-potassium pump is an example of an (active) transporter protein. The sodium-potassium pump exchanges sodium ions for potassium ions against their concentration gradient using ATP. This protein can be found in the plasma membrane. Cell Signaling Proteins: An example of a specific cell signaling protein is an adrenergic receptor (also a GPCR). Adrenergic receptors bind to ligands related to epinephrine, and its binding can stimulate the sympathetic nervous system. This protein can be found in the cell membrane. Genomic Caretaker Proteins: DNA topoisomerase is an example of a genomic caretaker protein. Its function is to regulate DNA supercoiling by catalyzing the winding and unwinding of the DNA strand(s). DNA topoisomerase can be found in the cell nucleus.
Within the aquaporin, each subunit contains a channel that allows H2O passage. Through which of the following views would you expect water to travel? Assume water is represented by the red dot. (IMAGE)
Picture C
What chemical properties of sodium dodecyl sulfate (SDS) contribute to equal charge/mass ratios needed for molecular mass determinations using SDS-PAGE? Why is the protein solution heat denatured prior to gel loading?
Sodium dodecyl sulfate (SDS) is an amphipathic molecule with negative charges that can be derived from its sulfate group. The significant negative charge of SDS contributes to equal charge/mass ratios needed for molecular mass determinations using SDS-PAGE by masking and giving proteins a net negative charge. The protein solution is heat denatured prior to gel loading to ensure that the mobility of the protein is not affected by its original structure and that the separation on the gel is actually based on a charge/mass ratio.
The enzyme active site provides an optimized physical and chemical microenvironment to lower the activation energy of a reaction and promote catalysis. Describe three catalytic reaction mechanisms used by enzymes and provide an example of each type using specific enzymes.
The first catalytic reaction is an acid-base catalysis reaction. This is a general reaction where a proton transfer involves a functional group. An example of this is pancreatic ribonuclease. This cuts RNA with assistance from histidine. The second catalysis reaction is covalent catalysis, and an example of this reaction is the formation of 1,3-bisphosphoglycerate from glyceraldehyde-3-phosphate thanks to the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase. The final catalytic reaction is metal ion catalysis. An example of this is carbonic anhydrase utilizing positive zinc ions to catalyze the reaction of bicarbonate derived from carbon dioxide.
What biochemical process must occur in the myosin head to initiate the "power stroke" in muscle contraction, which is characterized by a large protein conformational change? What conformational change is associated with ATP hydrolysis?
The myosin head has ADP + Pi at the nucleotide binding site in which Pi is released and induces the pull of actin filament to the center is in sarcomere, which is the biochemical process that must occur in the myosin head to initiate the "power stroke" in muscle contraction. The conformational change associated with ATP hydrolysis is the recovery conformational change. In this stage, the myosin head again has ADP and Pi waiting for Ca2+ binding to uncover the myosin binding sites on actin filaments.
What explains the 10,000 to 1 selectivity of the potassium channel for K+ over Na+ considering that the atomic radii of these two ions differ by only 0.4 Angstroms.
The selectivity channel, a narrow opening in the interior of the potassium channel protein complex, explains the 10,000 to 1 selectivity of the potassium channel for K+ over Na+. Both K+ and Na+ can go into the large chamber; however, only K+ ions can pass through the selectively channel and exit through the extracellular side of the membrane. The channel has amino acids that interact with the potassium in order to shed its hydration layers, but the sodium ions are smaller and therefore unable to interact with the amino acids.
The fractional saturation of hemoglobin (Hb) binding to oxygen is illustrated by A.) [HbO2] / [Hb]. B.) [HbO2] / ([HbO2] + [Hb]). C.) [Hb] / [HbO2]. D.) ([HbO2] + [Hb]) / [HbO2].
[HbO2] / ([HbO2] + [Hb]).
