You are trying to separate protein X from other proteins A and B in a solution.
ID: 151038 • Letter: Y
Question
You are trying to separate protein X from other proteins A and B in a solution. There
proteins have the following properties:
Protein
pI
Molecular weight (Da)
Protein A
8.5
90,000
Protein B
4.2
30,000
Protein X
8.5
30,000
What type of protein chromatography method might you use to separate protein X from protein A? Explain why. What type of protein chromatography method might you use to separate protein X from protein B? Explain why.
Protein
pI
Molecular weight (Da)
Protein A
8.5
90,000
Protein B
4.2
30,000
Protein X
8.5
30,000
Explanation / Answer
From the properties mentioned, we can see that protein A and protein X have same pI however differ greatly in molecular weight. Keeping this in mind, we can use size exclusion chromatograohy which is based on the size of the molecules, with the bigger protein eluting out first and the smaller protein eluting out later.
Also, what can be seen is that protein X and protein B have similar molecular weight but have different pI. This difference in pI can be targeted and we can use ion exchange chromatography to separate these proteins. As due to different pI, both the proteins will have a different charge (or intensity of charge) at different pH of the mobile phase (buffer). Based on this, the proteins' interaction with the stationary phase will be different and the two proteins can be eluted at different timings.
All the Best :)
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