There is a mutation in the lacI gene, called the Id mutation, which results in t

ID: 218864 • Letter: T

Question

There is a mutation in the lacI gene, called the Id mutation, which results in the lac operon being uninducible; that is, no ?-galactosidase is made in the presence or in the absence of lactose (these results are also unaffected by the presence or absence of glucose in the medium). In partial diploids (chromosome + plasmid), lacId acts as a dominant gene. In other words, even a good copy of the lacI gene on a plasmid cannot overcome the defect of the Id mutation on the genomic lacI gene.

a. (4 points) For what protein does the lacI gene code?

b. (4 points) To what region of DNA does the LacI protein bind?

c. (4 points) What is the likely cause of this phenotype?

d. (4 points) You have isolated a different mutant of E. coli (normal lacI gene) that has increased activity of the enzyme phosphodiesterase, which degrades cAMP. Predict the effect of this mutation on the inducibility of the lac operon.

e. (4 points) DNA footprinting using DNaseI revealed much information about how the lac repressor functions. What would DNA footprinting reveal about the DNA binding sites of a normal Lac repressor protein? A normal RNA polymerase?

Explanation / Answer

a. The reprossor protein is coded by lacI gene.

b. LacI protein binds to operator region of lac operon.

c. In the given case if the lacI gene can not synthesise a repressor protein , so the operator region can not be bound with the repressor, so the operator will be free always then it wont overlap with the promoter region and so all the lac operon will be switched 'on' always that is it will become cinstitutive operon.

d. cAMP is needed to form a complex with CAP which binds with the promoter region and makes the transcription high.When glucose is present cAMP is lowers down so no complex is formed and so no transcription occurs-that is called glucose effect.If cAMP gets degraded in that mutant then in presence of lactose also there will be little transcription of lac operon genes and proteins.

e. The lac repressor operates by a helix turn helix motif in its DNA binding domain binds to the base specific major groove of the operator region of the lac operon,with base contacts also made by residues of symmetry related alpha helices,'hinge' helices,which bind deeply in the minor groove.The DNA binding causes the specific affinity of RNA polymerases for the promoter sequence to increase sufficiently that it can not escape the promoter region and enter elongation,and so prevent transcription of the mRNA coding for the lac proteins.

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There is a mutation in the lacI gene, called the Id mutation, which results in t

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