5. Consider two proteins with a pl of 4.5 and pl. of 7.7? Using a cation-exchange column,describe how these two proteins could be separated. Indicate which buffer you wouldprepare and how a salt gradient may be employed.

Ion exchange chromatography (IEC) is a commonly used separation technique in biochemistry and…

Answered: Consider two proteins with a pl. of 4.5…

Curren'S Math For Meds: Dosages & Sol

11th Edition

ISBN:9781305143531

Author:CURREN

Publisher:CURREN

Chapter9: Parenteral Medication Labels And Dosage Calculation

Section: Chapter Questions

Problem 4.1P

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(E) Consider an amino acid that has one -amino group (pKa = 9.2), one -carboxyl group (pKa = 1.7) and one ionizable side chain (pKa = 6.2). At which pH range/s would this amino acid be effective as a buffer? If the amino acid shows a net charge of -1 at pH 11 and a net charge of +1 at pH 4, what pKa values would you use to calculate the isoelectric point? At what pH would the net charge be equal to zero?
Given a stock protein solution with a concentration of 6 mg/ml, determine the protein concentration of a solution made by mixing 5 μl of the stock with 5 μl of a buffer.
2 a. You are trying to purify protein C from a mixture of proteins noted in the above Table. If you had only one type of column to choose from, which one would allow you to purify protein C with the least number of contaminants? Size exclusion column Ion exchange column Affinity chromatography using glucose as the bait Affinity chromatography using NAD as the bait Please explain why you chose the column above based upon the properties of the column AND the proteins in the Table.
Three proteins with an overall charges of 0, -5, and -8 are passed through an anion exchange column equilibrated in a buffer with low ionic strength and eluted with an increasing gradient of salt in a buffer. Which protein will elute last from the column? 1. Protein with no charge. 2. Protein with eight negative charges. 3. Protein with five negative charges. 4. Both charged proteins will elute at the same time. O 2 O 4 O 3 O 1
Consider a crude extract with a mixture of the 5 proteins listed below. Protein A 4.5 pl 160 molecular weight, kDa Protein B 12.5 pl and 65 molecular weight Protein C 5.0 pl, 15 molecular weight Protein D 6.8 pl, 150 molecular weight Protein E 9.5 pl, 45 molecular weight You load this protein mixture onto an anion exchange column at pH 11. Next, you apply a "washing" step by passing through buffer at pH 11. Finally, for your elution step, you apply a pH gradient starting from pH 11 to pH 2.0 (A gradient buffer system allows you to gradually and continuously change the pH of your mobile phase starting from pH 11 up to pH 2). You load this same protein mixture onto a Size Exclusion column. Please indicate the order in which these proteins will elute for both. Group of answer choices Yes or No, please explain your answer. If your protein of interest is protein A, would using anion exchange column be completely successful at separating it from all the other proteins? Group of answer…
4. The table below describes a procedure in preparing standard solutions of a protein sample with different concentrations. (A) Determine the concentrations of each tube if the starting concentration of the protein is 500 mg ml (B) Determine the equation of the line if the concentration (x-axis) was plotted vs the absorbance (y-axis). (C) What is the concentration, in mg ml of the diluted unknown protein sample if the absorbance at 595 nm is 0.333? Tube Volume of protein, Volume of water, Final concentration, Absorbance mL mL mg ml [at 595 nm) 1 0.00 5.00 0.113 2 0.70 4.30 0.184 3 1.40 3.80 0.251 14 2.10 2.90 0.329 2.80 2.20 0.385 3.50 1.50 0.454 7 4.20 0.80 0.503 5.00 0.00 0.577
Discuss how buffers help prevent drastic swings in pH.
You had a second solution with an unknown concentration of Protein X that had to be diluted 4x before you could measure the absorbance. The diluted solution had an absorbance of 0.76. What is the concentration of the diluted solution in ug/ml and what is the concentration of the original solution
5 You need to separate the following three proteins, Protein A (MW 76 600 Da; pI 4.5); Protein B (MW 70 000 Da and pI 8.2) and Protein C (MW 42 000; pI 8.6). Describe what column/s you would use, the pH of the buffer/s you would use, which proteins are retained on the column and how you would elute the bound protein/s. How would you monitor where the protein peaks elute and whether the protein in each peak is pure of not?
6a. Using your 5 mg/mL stock solution, you wish to make 100µL of the correct concentration of protein to load. How do you make this working solution? (Yes- you would want this to be in 1x sample buffer, but that makes this question more complicated. So for this question just make the protein in water.) Volume of 10mg/ml Final Volume ! 1 100μL of protein 6b. This is a 1: A N Body* @ 2 W S dilution. #3 X E D $ 4 A C R F B I % 5 T V MacBook Pro U וכ H E U 8 N J ( 0
4. A solution containing egg albumin (pl-4.6), B-lactoglobulin (pl-5.2), and chymotrypsinogen (pl=9.5) was loaded onto a column of diethylaminoethyl cellulose (DEAE-cellulose) at pH 5.4.The column was then eluted with a pH 5.4 buffer, with an increasing salt concentration. Predict the elution pattern.
What amino acids will be positive at a pH of 2? Use the pKa table

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Consider two proteins with a pl. of 4.5 and pl. of 7.7? Using a cation-exchange column, describe how these two proteins could be separated. Indicate which buffer you would prepare and how a salt gradient may be employed.