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Exam Question 1 - ASFB 6101BCBMOL
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Question 1 of exam. Essay on protein purification, discussing each element and a justification for each, i.e. advantage of isothermal calorimetry, what it is.
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Student ID: 8298861. You have just purified a novel protein. Critically appraise different techniques to devise a
strategy you could use to identify and characterise the interactions of this protein, justifying
your final choices. Use diagrams as appropriate.
To decide which methods, I think are the most reliable and effective to identify and characterise the
novel protein I will discuss each method, the strengths, and weaknesses of each and explain why I
have chosen the methods I considered best for my investigation.
Firstly, I need to identify my protein, there are several techniques I could use. I could infer an
interaction with techniques such as genetic studies, bioinformatics, or cell perturbations, the two-
hybrid method, affinity chromatography, library surface display and in vivo studies.
st nd
1 Mutation 2 Mutation
Figure 1 – Double Surpressor mutation. A two-step process which a primary mutation to the gene
encoding the novel protein, disrupting the interaction between it and another molecule. Then a
second mutation allows the interaction to resume, suppressing the interaction disrupting effect of
the first. This method can be carried out using genetically amenable species such as Saccharomyces
cerevisiae.
st nd
1 Mutation 2 Mutation
Figure 2. Double enhancer mutation. The enhancer double mutation method involves a primary
mutation which has no effect on the interaction, a second mutation is made which disrupts the
interaction therefore enhancing the effect of the primary mutation. Saccharomyces cerevisiae is also
used for this technique.
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