BIO315 Molecular Biology Spring 2012
Dr. Henning Schneider
Study Guide T5
Regulation of Transcription inProkaryotes
1. The lac Operon

Concepts:

  1. The lac operon is a model systems for gene regulation in prokaryotes
  2. In negative control, an inducer interacts with a repressor to turn on gene expression
  3. Using mutants to study regulation of gene expression

Study Questions:

  1. Describe the organization of the lac operon.
  2. What effect has lactose on the expression of the beta-galactosidase gene?
  3. What is the difference between a single and a double operon?
  4. Explain the experiment that demonstrates the binding of the repressor to DNA in the presence or absence of the inducer.
  5. Explain the experiment that shows binding of the repressor to DNA in operator-constitutive mutants.
  6. Explain why mutations can occur in different genes and regions of genes and still have the same phenotype?
  7. Explain the role of the repressor in preventing RNAP to enter the elongation phase in lac operons?
  8. Explain the term "negative control".
  9. Explain the negative control mechanism of the lac operon.
  10. Why is it important that the repressor forms a tetramer complex?
  11. What are merodiploid bacteria?
  12. Explain the term constitutive mutant.
  13. How does a mutation of the repressor gene lead to a constitutive mutant?
  14. Explain why a mutation of the lac repressor gene can be recessive.
  15. Explain why a mutation of the lac repressor gene can be dominant.
  16. Give an example of a cis-dominant mutation.
  17. Give examples of dominant mutations that lead to constant gene activity or constant gene repression.
  18. Did cis-mutations prove the existence of an operator? Why?
  19. What effect has a mutation of the repressor that cannot recognize the operator?
  20. Why are all three lac genes (z, y, and a) effected by a mutation in the operator?
  21. Why are mutations in the repressor gene more likely to be dominant than a mutation in the operator?
  22. Explain the experiment in which the binding of the RNAP to the promoter in the presence of the repressor was demonstrated.
  23. Explain how a nitrocellulose filter binding assay works.

2.The lac operon: positive control mechanisms

Concepts:

  1. Positve control of an operon
  2. Catabolite Activator Protein (CAP)
  3. CAP-DNA interactions
  4. CAP-RNAP interactions

Study Questions:

  1. Define the term positive control of an operon.
  2. Explain the function of additional operator region upstream and downstream of the major operator at the transcription initation start site.
  3. Describe the arrangement of all three operators and the CAP site of the lac operon.
  4. Explain the mutation experiment (Müller-Hill et al.) that demonstrates the repression efficiency of all lac operators.
  5. Explain the function of the catabolite activating protein.
  6. What effect has cAMP on the activity of the lac operon?
  7. Explain the experiment that demonstrates the effect of CAP on beta-galactosidase activity.
  8. Explain the mechanism of CAP action.
  9. Which subunits of the RNAP interact with CAP at the promoter?
  10. Describe the interaction of CAP with cAMP and DNA.
3. The ara operon

Concepts:

  1. Catabolites such as arabinose autoregulate the biosynthetic pathway.
  2. Multiple operators in one operon are associated with looping of DNA.
  3. The ara operon is controlled by looping of DNA.
  4. Looping of DNA in the ara operon is based on interaction of operator elements, CAP, repressor, and inducer.

Study Questions:

  1. Describe the arrangement of regulatory sites of the ara operon.
  2. Explain the function of the ara C gene product.
  3. Explain the control/autocontrol of the araC gene.
  4. Describe the organization of the ara repression loop.
  5. What is the function of arabinose in the control of the ara operon?
  6. Describe the experiment that demonstrates looping of DNA by the araO2 and araI regions.
  7. Which method was used to distinguish between and looped and unlooped form of the operon?
  8. What protein interactions are critical for the regulation of the ara operon? Which proteins bind toeach other? Which bind to DNA? Which changes occur when transcription is turned on and turned off?
  9. Which conclusions can be drawn from the looping experiment?
  10. Explain the concept of supercoiled DNA.
  11. Identify positive and negative control mechanisms in the ara operon.
  12. What was the role of araO2 and araI mutants in experiments that established the regulatory mechanism in the ara operon?
  13. Which interactions lead to the breaking of the loop in the ara operon?
  14. Which experiments demonstrated that the loop formation in the ara operon is reversible? Explain the experiment.
  15. What is the function of araI1 and araI2 elements in the ara operon?
  16. How the interaction of araI2 and araI1 elements in the ara operon was demonstrated experimentally.
4. The trp operon

Concepts:

  1. Tryptophan has a negative role in controlling the trp operon

Study Questions:

  1. Explain the function of the trp operon.
  2. Describe the arrangement of control elements on the trp operon.
  3. Explain the control of the trp operon through the trp repressor.
  4. What is the function of the tryptophan in the control of the repressor?
  5. What type or operon is the trp operon?

Last modified on April 16, 2012 by HS.