Some characteristics of three proteins are listed in the table below: Protein Mo
ID: 173873 • Letter: S
Question
Some characteristics of three proteins are listed in the table below:
Protein
Molecular Weight (Da)
Isoelectric point (pI)
Does the Protein Contain a heme moiety?
1
15,000
10.5
No
2
14,000
5.0
No
3
65,500
4.5
Yes
a.(2 points) What type of chromatography separates proteins based on size?
b. (2 points) What type of chromatography separates proteins based on their charge?
c.(10 points) Could gel filtration chromatography be used to separate a mixture containing Protein 1 and 3? Clearly explain why or why not. If gel filtration chromatography can be used to separate Protein 1 from Protein 3, which protein would elute first (clearly explain why)? After collecting the fractions from the column, the absorbance of each fraction will be measured using a spectrophotometer. Can both proteins (Protein 1 and Protein 3) be monitored at 280nm and 400nm (clearly explain)?
d. (6 points) Which 2 proteins listed in the table above could be separated by ion exchange chromatography but not by gel filtration chromatography (clearly explain why)?
Protein
Molecular Weight (Da)
Isoelectric point (pI)
Does the Protein Contain a heme moiety?
1
15,000
10.5
No
2
14,000
5.0
No
3
65,500
4.5
Yes
Explanation / Answer
A-Separation on the basis of Size -Gel Filtration Chromatography
B-Separation on the basis of their charges-Ion Exchange and Chromatofocusing Chromatography
C_ Gel Filtration Chromatograpgy is a technoques based on the separation of the proteins based on size. 65000 Da will be eluted first and then the 15000 Da. Proteins with heme moeity will show absorbance at 280nm will without heme moeity will show absorbance at 400nm.
D-proteind 2 na d3 will easuily be separated by the ion exchange chtromatography.
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