(a) You decide to purify the 5-HT 2A receptor. The protein has a predicted molecularMass of 46.54 kDa and isoelectric point of 8.94. Will the5-HT2A Receptor proteinBind to an anion exchange column if you are using a buffer containing 150m M NaCl, 50mM carbonate buffer, pH6.0, and 10% glycerol?(b) After performing affinity chromatography purification, you run a SDS page gel showingThe following:25015010075503725.1510StandardsSample| || |Explain why you see four bands in the sample lane? Label the band that corresponds toYour protein of interest. What do you think is going on here? (c) You decide to run an additional purification step using size exclusion chromatography.Draw what you expect to see on the resulting chromatograph.A₂80(d) You run another SDS page gel. Assuming your purification was successful, draw the resultYou expect to see in the sample lane.2501503010075503725Elution volume (ml)1510StandardsSample

Formation of the peptide bond neutralises any charge due carboxyl group, therefore the net charge on…

Answered: (a) You decide to purify the 5-HT 2A…

Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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) Dr Merrick is so excited to know the affinity constant Kd values for O2 binding to mammoth OBP and elephant hemoglobin at pH=7, that he starts speculating about the kinetics of binding to O2. If the ka value of mammoth EHb-O2 complex was the same as the ka value of mammoth OBP-O2 complex, what would be the ratio between the kd values of these complexes? Explain your reasoning. Hint: we are looking for the ratio kd(EHb)/kd(OBP) at pH=7.
The order of elution of amino acids X, Y, VW and Z in a cation exchanger eluted with a buffer of increasing pH is as follows: Z, W, Y, X (last). If the amino acids are glutamic acid, arginine, phenylalanine and alanine, what amino acid can Z be? O A O F O E OR
A new oxygen transport protein that exhibits cooperative binding has been isolated and is beingstudied in the lab. Calculate the KD value if Y = 0.76 when pO2 = 18 torr (assume n = 2.5). Howdoes this compare to the KD value for hemoglobin? Does this protein bind more or less tightly tooxygen compared to hemoglobin?
In an experiment, hemoglobin is dissociated in a buffer and a subunit is isolated to study for its oxygen binding affinity. (i) What is the shape of the oxygen dissociation curve is expected in the experiment?Explain why. (ii) Is the Km of the isolated subunit higher or lower than the Km of an intact hemoglobin?
An experiment was carried out to measure the reaction rate of hydrolysis of acetylcholme (substrate) with serum enzymes (Eadie, 1949). In the experiment, two experiments were conducted, namely experiment 1 without using a prostigmine inhibitor and experiment 2 using a prostigmine inhibitor at 1.5 x 10^-7 mol/l. the data obtained are: a. Is prostigmine competitive or noncompetitive inhibitor? b. determine the value of km and rmax for the two experiments, compare
Part B Calculate the logarithm (log) of the concentration of calcium in the cytosol. Express your answer numerically to one decimal place. ΑΣφ ? Submit Request Answer Part C Convert the concentration of calcium in the blood to scientific notation. Complete the value by entering a coefficient and an exponent numerically.
Acetic acid (a weak acid with a pKa of 4.75) and ethanol (an alcohol) are each composed of two carbons, hydrogen, and oxygen, and both enter cells by passive diffusion. At pH 7, one is much more able to permeate a cellular membrane than the other. Which is more membrane permeable, and why? Predict how the membrane permeability of each is altered when the extracellular pH is reduced to 1.0, a value typical of the stomach.
An oligopeptide has the following amino acid sequence: NH2-Ala-Glu–Leu–Trp–Tyr-Ser–Gly–Lys–Leu-Ala–Arg-Ala-Phe-Ile-Pro–Gly-COOH a) Estimate the net electric charge of the molecule at pH 8.0 and pH 11.0. b) If the above peptide is passed through a cation exchange chromatographic column (that is, the matrix of the column has negative charges) stabilized at pH 8.0, would you expect it to be retained on the column? c) Indicates the number of fragments, and the sequence of each of them, that would be obtained when treating the peptide in question with: i) trypsin, ii) chymotrypsin.
An antibody binds to another protein with anequilibrium constant, K, of 5 × 109 M–1. When it binds toa second, related protein, it forms three fewer hydrogenbonds, reducing its binding affinity by 11.9 kJ/mole. Whatis the K for its binding to the second protein? (Free-energychange is related to the equilibrium constant by the equa-tion ΔG° = –2.3 RT log K, where R is 8.3 × 10–3 kJ/(mole K)and T is 310 K.)
The equilibrium constant for the hydrolysis of the peptide alanylglycine (Gly-Ala in the reaction from Part B) by a peptidase is K = 9.0 × 10² at 310 K. Calculate AG for this reaction. Express the Gibbs free energy to three significant figures. AG = Submit ΠΑΠΙ ΑΣΦ Request Answer ? kJ/mol Keq [Gly] [Ala] [Gly-Ala]
Draw a schematic of the chemical structure of the Leu-Tyr-Met-Lys tetrapeptide, with the correct ionization at neutral pH (pH 7). Also, give an estimate the pl of this tetrapeptide (you are given that the pKa of any acidic R group is 4, the pKa of any basic R group is 11.5, and the pKa of the α-COOH is 2, and the a-NH3 is 9.5). Provide rationale!
The Graph below shows the binding curves of two proteins (A and B) for the same ligand (L). Use this Graph and determine the dissociation constant, K, for both proteins. Which protein (A or B) has a greater affinity for ligand L? Which of the two proteins would be more easily inhibited by an antagonist? 1.0 Y 0.5 2 A 4 6 B 8 [L] (μM) 10 12 14 16

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