Question

Question 11 [10 marks / 10 minutes]
For the mutations in the 5’ UTR of the trp operon given below, explain the effect on expression of the trp operon relative to wildtype (increased, decreased, or unchanged).
Assume medium levels of tryptophan, such that TrpR (repressor) is inactive and attenuation is active.
a) A mutation in region 2 such that it cannot bind with region 3. [4 marks]
b) A mutation in region 4 such that it cannot bind with region 3. [4 marks]
c) Insertion of T-A basepairs immediately after region 3, such that uracils are transcribed. This insertion does not change the stem-loop formation properties of regions 1, 2, 3 or 4. [2 marks]

Answer 1

The trp operon is a repressible operon which means that the gene expression decreases in presence of the substrate for the operon. Here, the interaction of the substrate with the operon induces a physical change in the structural organization of the operon which prevents linear passage of the RNA polymerase and hence prevents gene transcription. Thus, if a mutation results in 5' UTR region, the consensus and conserved sequence of the gene promoter would be deteriorated and RNA polymerase would not be able to find and recognize it. This will lead to decrease or altogether failure of gene expression.

Answer a: Increase (At medium levels of tryptophan, the regions 2 and 3 tend to make the stem-loop structure but owing to mutation, they fail to do so. Hence, the RNA polymerase can traverse through the operon swiftly. Considering that attenuation is active, the RNA polymerase would fail to traverse and gene expression would increase)

Answer b: Unchanged (At medium levels of tryptophan, the regions 2 and 3 tend to bind at the place of regions 3 adn 4. Hence, the gene expression levels would remain unchanged)

Answer c: Unchanged (Since the stem-loop formation capability remains unchanged even after the mutation, the rate and extent of gene expression would remain unchanged)