Topic 5: Protein Purification Techniques
Isolation and analysis methods, chromatography, electrophoresis.
Cell Disruption Techniques Identification
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Which one of the following cell disruption techniques correspond to the description 'Cells are suspended in buffers containing protein catalysts that digest the cell wall.'.
Types of Column Chromatography
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Which one of the following types of column chromatography correspond to the description 'separates molecules by differences in size as they pass through a porous particles in a column'.
Identification of Macromolecules in Gel Electrophoresis Processes
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Which one of the following types of macromolecules correspond to the gel component or process 'electrical fields'.
Determining Net Charge of Proteins at Given pH
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Isoelectric Point Problem
| Protein Name | isoelectric point (pI) | molecular weight |
|---|---|---|
| Transferrin (Tran) | 5.9 | 80.0 |
The protein in the table (above) is placed in a buffer solution with a pH of 4.5.
What is the correct net charge on the Tran protein at pH of 4.5
Protein Migration Direction Based on Isoelectric Point
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Isoelectric Point Problem
A mixture of two proteins are to be separated by isoelectric focusing.
| Protein Name |
Isoelectric Point (pI) |
Molecular Weight |
|---|---|---|
| Horseradish peroxidase (HP) | 9.0 | 34.0 |
| α-Amylase (Amy) | 5.9 | 61.0 |
Both protein samples are placed into a gel with a constant pH of 7.5. The gel is then placed into an electric field.
In which direction will each protein in the table migrate at pH 7.5?
Estimating Protein Molecular Weight from SDS-PAGE Migration
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Gel Migration Problem
In this task, data from an SDS-PAGE experiment, where proteins are separated based on molecular weight, is provided. The gel results table below shows some standard proteins with known molecular weights and one unknown protein.
| Protein Name | Molecular Weight (kDa) |
Migration Distance (cm) |
|---|---|---|
| Streptavidin (Stp) | 13.2 | 3.44 |
| Avidin (Av) | 16.9 | 3.24 |
| Trypsin (Tryp) | 23.5 | 2.97 |
| Elastase II (Ela) | 26.5 | 2.88 |
| Lipase (Lip) | 40.0 | 2.55 |
| Luciferase (Luc) | 50.0 | 2.37 |
| Catalase (Cat) | 65.5 | 2.15 |
| Unknown | ? | 3.37 |
Estimate the molecular weight of the unknown protein by comparing its gel migration distance with those of the standards.