4. IPTG is an analog of lactose which is used to release the repressor on the lac operon allowing our protein to be produced. a. Draw IPTG and lactose and indicate the portions that are the same for both structures. b. Propose a reason that IPTG is used instead of simply adding lactose.
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- Answer as Directed. Below is the model of a lac operon. lac I lac Z с promoter operator lac Y lac A DNA +1 1. What are structural genes? Are the lac structural genes transcribed in the absence of lactose? 2. What is the role of the promoter and operator sites in the operon? 3. Is the repressor protein bound to the operator site in the absence of lactose? In its absence? 4. Under what nutritional circumstances (high or low glucose) is CAP bound to cAMP? 5. In the absence of lactose and the presence of glucose in the bacterial growth media, what proteins are bound to the lac control region? Is the operon being transcribed then? 6. In the presence of lactose and the presence of glucose in the bacterial growth media, what proteins are bound to the lac regulatory region? Is the operon being transcribed then? 7. In the presence of lactose and the absence of glucose in the bacterial growth media, what proteins are bound to the lac control region? 8. Why is it adaptive for a bacterium to not…. Recall that the trp operon has a special leader sequence (trpL) between the operator and the structural genes that offers attenuation as a mechanism for regulation of gene expression. (A) Draw a diagram of a trpL region of the operon when tryptophan is abundant in the cell.Label the following features: the DNA, 5’ and 3’ polarity of the RNA, the regions 1, 2, 3,and 4 and poly-U of the RNA, the pair of Trp codons (UGG), the ribosome, and RNA-Pol,along with any stem-loop structure that would form under these conditions (B) In the above example, will the rest of the trp operon genes be expressed? Briefly describe your reasoning why or why not (C) The trp codons in region 1 of the trpL gene have mutated to cysteines (UGG to UGC). What will be the effect on attenuation gene regulation of the trp operon? Brieflyexplain your reasoning.Regulation of Genes and Their products 1. Given the following genotypes, explain how the mutation (identified by a (-) superscript) wil affect E. coll grown in lactose medium. Will the lac operon be on or off? Will there be a complete set of gene products from the lac operon? What will be the implication of the missing gene product, if ever? Will the cell be able to survive in the lactose medium or not? a. I+p+o+z- y+ b. i- p+o+z+y+ c. i+p+o- z+y+ d. i+p- o+z+y+ 2. In terms of the trp operon, differentiate between two normal bacterial cultures, one grown in a medium supplied with tryptophan and the other medium without tryptophan. 3. Experiments show that mutations at gene E lead to non-repressible transcription of trp genes. Why?
- Read aloud V Draw Highlight 2. You are studying the regulation of the lactose operon in Escherichia coli, by measuring expression of the lacZ gene (i.e production of beta-galactosidase). (a) You identify several loss-of-function mutations in which lacZ is never expressed, in the presence and absence of glucose and lactose. What components of the lac operon could be mutated to produce this phenotype? List all possibilities. (b) You identify another loss-of-function mutation with the following expression pattern: Media + glucose - lactose + glucose +lactose - glucose - lactose - glucose + lactose lacZ expression Low Low High High What components of the lac operon could be mutated to produce this phenotype? List all possibilities.Answer as Directed. Below is the model of a lac operon. lac I lac Z с promoter operator +1 lac Y lac A DNA 1. In the absence of lactose and the presence of glucose in the bacterial growth media, what proteins are bound to the lac control region? Is the operon being transcribed then? 2. In the presence of lactose and the presence of glucose in the bacterial growth media, what proteins are bound to the lac regulatory region? Is the operon being transcribed then? 3. In the presence of lactose and the absence of glucose in the bacterial growth media, what proteins are bound to the lac control region? 4. Why is it adaptive for a bacterium to not express the genes that encode for that lactose utilization proteins when lactose is not available or when glucose is present? 5. Why is it adaptive for the structural genes for using lactose to be under the control of a single promoter, i.e., synthesize a polycistronic message rather than three monocistronic message?. An interesting mutation in lacI results in repressorswith 110-fold increased binding to both operator andnonoperator DNA. These repressors display a “reverse”induction curve, allowing β-galactosidase synthesis inthe absence of an inducer (IPTG) but partly repressingβ-galactosidase expression in the presence of IPTG. Howcan you explain this? (Note that, when IPTG binds a repressor, it does not completely destroy operator affinity,but rather it reduces affinity 110-fold. Additionally, ascells divide and new operators are generated by thesynthesis of daughter strands, the repressor must findthe new operators by searching along the DNA, rapidlybinding to nonoperator sequences and dissociating fromthem.)
- protein. You create a mouse line with Cas9 under control of a brain-specific enhancer, while the short guide RNA complementary to the first exon of Gene Y is expressed in all tissues. You subsequently sequence Gene Y in both brain and liver tissue. What would expect in each tissue? You can assume that the CRISPRICas9 system will impact both copies of Gene Y in cells, and that the first exon of Gene Y is necessary for Gene Ys function. a. Liver: Functional Gene Y; Brain: Functional Gene Y b. Liver: Nonfunctional Gene Y; Brain: Funtional Gene Y c. Liver: Functional Gene Y; Brain: Nonfunctional Gene Y d. Liver: Nonfunctional Gene Y; Brain: Nonfunctional Gene YAll about splicing A. snRNPs interactions will bring the 5' and 3' splice sites together in the pre-mRNA B. lariat formation is necessary to bring the branch site with the 5' splice site of the intron C. an unstable 5'P and 2'OH phosphodiester bond helps form the lariat D. exon 5' splice site consensus sequence GU and its 3' splice site AG are recognized by snRNPs E. various types of snRNPs and the pre-RNA come together to form the spliceosome B, C, D, E only A, B, C, E only A, B, C, D, E A, B, C onlyGTTTTCACTGGCGAGCGTCATCTTCCTACT 8. What is the function (e.g. transcriptional regulation, transmembrane signaling, kinase, protease, etc.) of the protein(s) encoded by the gene.
- All about splicing A. snRNPs interactions will bring the 5' and 3' splice sites together in the pre-mRNA B. lariat formation is necessary to bring the branch site with the 5' splice site of the intron C. an unstable 5'P and 2'OH phosphodiester bond helps form the lariat D. exon 5' splice site consensus sequence GU and its 3' splice site AG are recognized by snRNPs E. various types of snRNPs and the pre-RNA come together to form the spliceosome A, B, C, E only A, B, C, D, E A, B, C only B, C, D, E only. Why is a nonsense suppressor tRNATyr, even though ithas a mutant anticodon that cannot recognize a tyrosinecodon, charged with tyrosine by Tyr tRNA synthetase?. CTP synthetase catalyzes the glutamine-dependent conversion of UTP to CTP. The enzyme is allosterically inhibited by the product, CTP. Mammalian cells defective in this allosteric inhibition are found to have a complex phenotype: They require thymidine in the growth medium, they have unbalanced nucleotide pools, and they have a mutator phenotype. Explain the basis for these observations.
