1) E. coli cells are growing in medium containing lactose but no glucose. Briefly describe the consequence of the following changes (one to two sentences for each condition):
a. Addition of high concentration of glucose. b. A mutation that inactivates galactoside permease. c. A mutation that inactivates beta-galactosidase. d. A mutation that affects binding of CAP to c-AMP e. A mutation that affects binding of inducer to LacI f. A lac operator mutation that deletes all of the O1
2) Describe the allosteric regulation model for transcription termination and briefly indicate how this model can explain both Rho dependent and Rho independent termination reactions.
3)Please provide brief (one sentence) answers:
a) The Mycoplasma chromosome codes for 500 genes, yet it has about 20 promoters. Provide a brief explanation for how this is possible.
b) How many seconds would it take the bacterial RNA polymerase to synthesize an mRNA of 5,000 bases?
c) How many mistakes would the bacterial RNA polymerase make while synthesizing an mRNA of 30,000 bases?
d) Sigma (?) protein by itself does not bind to promoter sites. Predict the effect on transcription of a mutation enabling ? to bind tightly to the –35 region in the absence of other subunits of RNA polymerase.
e) What is the structure of the most common transcription terminator signal in E. coli, and what are 2 conserved features of the RNA sequence of this structure?
f) Why is RNA synthesis not as carefully monitored for errors, as is DNA synthesis?
4) You started work in a transcription lab working a with a novel bacteria phage. You isolated an enzyme that has RNA polymerase activity, and you discovered that your new viral RNA polymerase has a unique property. Unlike E. coli RNA polymerase, which prefers starting transcription with ATP, your new viral polymerase prefers starting transcription with GMP. You know the directionality of RNA chain growth (3’-to-5’ or 5-to-3’) by your new RNAP.
A. Explain in what direction (3’-to-5’ or 5-to-3’) this enzyme synthesizes RNA and how you know this.
B. What other simple experiment can you do to show that your conclusion is correct, provided you have full access to all rNTPs, 3’-dNTPs, 2’-dNTPs?
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