Biochemistry: Concepts and Connections (2nd Edition)
2nd Edition
ISBN: 9780134641621
Author: Dean R. Appling, Spencer J. Anthony-Cahill, Christopher K. Mathews
Publisher: PEARSON
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Chapter 6, Problem 13P
A protein gives under conditions of buffer composition, pH, and temperature that are close to physiological conditions, a molecular weight by size exclusion measurements of 140,000 g/mol. When the same protein studied by SDS gel electrophoresis in the absence or presence of the reducing agent
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A protein gives, under conditions of buffer composition, pH, and temperature that are close to physiological conditions, a molecular weight by sedimentation equilibrium measurements of 140,000 g/mol. When the same protein is studied by SDS gel electrophoresis in the absence or presence of the reducing agent β-mercaptoethanol (BME), the patterns seen in lanes A and B respectively are observed. Lane C contains standards of molecular weight indicated. From these data, describe the native protein, in terms of the kinds of subunits present, the stoichiometry of subunits (that is, how many of each subunits are present), and the kinds of bonding (covalent, non-covalent) existing between subunits
. A protein gives, under conditions of buffer composition, pH, and tem-
perature that are close to physiological conditions, a molecular weight
by sedimentation equilibrium measurements of 140,000 g/mol. When
the same protein is studied by SDS gel electrophoresis in the absence
or presence of the reducing agent B-mercaptoethanol (BME), the pat-
terns seen, respectively, in lanes A and B are observed. Lane C contains
standards of molecular weight indicated. From these data, describe the
native protein, in terms of the kinds of subunits present, the stoi-
chiometry of subunits, and the kinds of bonding (covalent, noncova-
lent) existing between subunits. /
A
B
- BME
+ BME
STD
Serum albumin,
67,000
Ovalbumin,
43,000
Carbonic anhydrase,
30,000
Trypsin inhibitor,
20,000
10
2.
4.
Co
cm migrated
suctose density gradient ultractifugation is a powerful technique for fractionating macromolecules like DNA,RNA and proteins.some protocols using sucrose gradients mention the following:10 mL sucrose gradients 10-15%(w/v) in 10 mM HEPES buffer.How would you prepare this sucros gradient?
Chapter 6 Solutions
Biochemistry: Concepts and Connections (2nd Edition)
Ch. 6 - Prob. 1PCh. 6 - Bovine pancreatic trypsin inhibitor (BPTI; Figure...Ch. 6 - A schematic structure of the subunit of...Ch. 6 - In the protein adenylate kinase, the C-terminal...Ch. 6 - Give two reasons to explain why a proline residue...Ch. 6 - Consider a small protein containing 101 amino acid...Ch. 6 - a. Based on a more conservative answer to Problem...Ch. 6 - The following sequence is part of a globular...Ch. 6 - a. A protein is found to be a tetramer of...Ch. 6 - Under physiological conditions, the protein...
Ch. 6 - Theoretical and experimental measurements show...Ch. 6 - The peptide hormone vasopressin is used in the...Ch. 6 - A protein gives under conditions of buffer...Ch. 6 - A protein gives a single band on SDS get...Ch. 6 - It has been postulated that the normal...Ch. 6 - Below are shown two views of the backbone...Ch. 6 - Do you expect a Pro Gly mutation in a...Ch. 6 - Rank the following in terms of predicted rates...Ch. 6 - Shown below are two cartoon views of the small...Ch. 6 - Prob. 20PCh. 6 - In most cases, mutations in the core of protein...Ch. 6 - A Leu Ala mutation at a site buried the core of...Ch. 6 - Disulfide bonds have been shown to stabilize...Ch. 6 - Cartoon renderings of the proteins Top 7 and adaH2...
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- The A280 of a protein sample loaded onto a gel was determined to be 0.767 (1.00 cm path length, after subtracting the blank). The total volume of this sample was 428 µL. 19.0 µL of this protein sample was mixed with 19.0 µL of 2X laemalli sample buffer and then 12.0 µL of the entire sample was loaded into the gel and electrophoresed. Calculate the amount of protein that was loaded into the gel (in µg).arrow_forwardThe protein concentration of a known standard is 100mg/mL If you prepared a serial dilution, mixing 10μL of protein with 40μL of water what would be concentrations of the first 3 dilutions?arrow_forwardcalculate the volume of stock solutions required to make up the buffer solutions that will be used for protein purification. The solutions you need to prepare for purification are: i. Binding Solution A: make up 50 mL 50 mM HEPES buffer (pH 7.5), 300 mM NaCl, 5mM imidazole, 5% (v/v) glycerol ii. Wash Solution B: make up 50 mL 50 mM HEPES buffer (pH 7.5), 300 mM NaCl, 75mM imidazole, 5% (v/v) glycerol iii. Elution Solution C: make up 10 mL 50 mM HEPES buffer (pH 7.5), 300 mM NaCl, 500 mM imidazole, 5% (v/v) glycerol please show your working . Thnk youarrow_forward
- 80mL of a 0.3M solution of hexapeptide Leu-His-Cys-Glu-Asn-Arg is adjusted to pH=pl. The solution is then titrated with 0.2M HCI to a final pH of 2.1. Sketch the titration curve, labelling the pH and volume axes. Indicate the volume of HCl needed to reach each relevant pKa value and equivalence point(s). Relevant pka values are: 2.1, 4.3, 6.0, 8.3, 9.8, and 12.5.arrow_forwardProtein concentration can readily be determined using the Beer-Lambert law: A = e l c where A = absorbance e = molar absorption coefficient (M-1cm-1) l = light path length (cm) c = concentration (M) If the molar absorption coefficient at 280 nm for yeast ADH is 48860 M-1cm-1 and a 10 mL solution of the protein has an absorbance at 280 nm of 0.4 (as measured by a spectrometer with pathlength 1 cm), then what is the concentration of the protein solution (in μM)? i.e. concentration = ______ μM If the molecular weight of the protein is 36849, what is its concentration in mg/mL? i.e. concentration = _______ mg/mL For each part of the question, show your calculations to arrive at your answers.arrow_forwardThe diffusivity of amino acids in polyacrylamide gel is approximately 1x10^-9 cm2./s calculate the initial flux of amino acids, give an instantaneous gradient of (20g/cm 3 )/8cm Why is polyacrylamide gel is used in electrophoresis?arrow_forward
- Consider the following properties of the protein components of a sample mixture as provided in the table below: 1. if the mixture is subjected to gel filtration chromotography which protein component elute first? 2. if the mixture is subjected to isoelectric focusing which protein will stop m oving nearest to the positive electrode? 3. if the mixture is subjected to cation-exchange chromotography using a buffer at ph 7 which protein will bind to the resin? 4.if the mixture is subjected to SDS-PAGE which protein will be at bottomost portion of gel? 5.if the mixture is subjected to hydrophobic interaction chromotography which protein will bind most strongly to the resin?arrow_forwardIn RP HPLC, one of the most common separation methods used to measure purity, strength, dosage, etc, a protein would be put into 0.1% (1000 ppm) TFA (Trifluoroacetic acid), what do you suppose this does to the protein in many cases? (Pick the BEST answer). somewhat denaturing to very denaturing oxidizes cysteine ionizes acid and base groupsarrow_forwardDiethylaminoethyl cellulose is a positively charged resin used in ion-exchange chromatography with a pKa is about 11. A negatively charged protein with an isoelectric point of 5.0 is applied to the column in a buffer at pH 7 and containing 0.1 M NaCl. Which of the following conditions is most likely to weaken the interaction between the protein and the resin? A Raising the pH to 8 and decreasing the NaCl to 0.05 M B Raising the pH to 8 and increasing the NaCl to 0.2 M C Lowering the pH to 6 and decreasing the NaCl to 0.05 M D Lowering the pH to 6 and increasing the NaCl to 0.2 Marrow_forward
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