PRESCOTT'S MICROBIOLOGY
11th Edition
ISBN: 2818440045677
Author: WILLEY
Publisher: MCG
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Chapter 18, Problem 2AL
Summary Introduction
EMSA stands for electrophoretic mobility shift assay and it is the most common method used to study DNA-protein interactions. This method is also termed as a gel shift assay. This is because the DNA binding protein is added to the purified target DNA mixture and its mobility is slowed down using an agarose gel. The DNA-protein complex is found to be shifted to a higher molecular weight when compared to the DNA without the protein. Usually, the magnitude of the shift is related to the DNA-protein ratio. Thus, the largest shift would be seen when sufficient protein is added.
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The lac operon of E. coli controls the expression of genes that code for enzymes involved in lactose metabolism. Mutations to various regions of the lac operon have been discovered which affect both the control of gene expression and the function of the gene products. These mutations include:
I - = produces a repressor protein that cannot bind to the operator region
IS = produces a repressor protein that cannot be removed from the operator region
OC = cannot bind a repressor protein
Z - = produces a defective (non-functioning) β-galactosidase enzyme
+ = wild type
Detail the functionality of the lac operon under the following genetic and cellular conditions. You do not need to consider the role of CAP in your analysis.
Genes
Repressed (yes/no)
Constitutive (yes/no)
I+ OC Z+
I+ O+ Z+
(Lactose
present)
I- O+ Z+
The lac operon of E. coli controls the expression of genes that code for enzymes involved in lactose metabolism. Mutations to various regions of the lac operon have been discovered which affect both the control of gene expression and the function of the gene products. These mutations include:
I - = produces a repressor protein that cannot bind to the operator region
IS = produces a repressor protein that cannot be removed from the operator region
OC = cannot bind a repressor protein
Z - = produces a defective (non-functioning) β-galactosidase enzyme
+ = wild type
Detail the functionality of the lac operon under the following genetic and cellular conditions. You do not need to consider the role of CAP in your analysis.
Genes
Repressed (yes/no)
Constitutive (yes/no)
IS O+ Z+
IS OC Z+
The amino acid asparagine is synthesized from aspartic acid by the enzyme asparagine synthetase (AS).
In the previous problem you proposed a model for how this gene could be regulated. Suppose that you carry out an experiment to test your model. To do this you cut out the regulatory sequences upstream of the gene and fuse it to a gene for green fluorescent protein (GFP). Now you can visually observe when the gene is activated. You insert this engineered gene into a host cell and look for GFP expression. You discover some mutants that have different expression levels of GFP and call them GFP1- and GFP2-. The expression levels of GFP are given below.
Cell
GFP expression
Wild type
100
GFP1-
50
GFP2-
0
Propose an explanation for these results based on your model. In other words, what was mutated and how?
This answer should include whether the mutation is (view links for more information):
dominant or recessive https://www.ncbi.nlm.nih.gov/books/NBK21578/#A1877
in a cis…
Chapter 18 Solutions
PRESCOTT'S MICROBIOLOGY
Ch. 18.1 - MICRO INQUIRY What is the function of the 3-OH...Ch. 18.1 - MICRO INQUIRY Why is it important that identical...Ch. 18.2 - MICRO INQUIRY Which step (or steps) in this...Ch. 18.2 - Retrieve, Infer, Apply Why is the Sanger technique...Ch. 18.2 - Retrieve, Infer, Apply Explain the difference...Ch. 18.2 - Retrieve, Infer, Apply Why does reversible chain...Ch. 18.2 - Prob. 4CCCh. 18.2 - Retrieve, Infer, Apply Suggest a medical and an...Ch. 18.3 - Retrieve, Infer, Apply NGS techniques are...Ch. 18.3 - Retrieve, Infer, Apply Examine figure 18.8. How...
Ch. 18.4 - Prob. 1MICh. 18.4 - Prob. 1CCCh. 18.4 - Prob. 2CCCh. 18.4 - Prob. 3CCCh. 18.5 - Figure 18.12 Metabolic Pathways and Transport...Ch. 18.5 - Prob. 2MICh. 18.5 - Prob. 3MICh. 18.5 - Prob. 1CCCh. 18.5 - Retrieve, Infer, Apply How might the following...Ch. 18.5 - Retrieve, Infer, Apply Compare and contrast...Ch. 18.5 - Retrieve, Infer, Apply Why does two-dimensional...Ch. 18.5 - Retrieve, Infer, Apply What is the difference...Ch. 18.5 - Retrieve, Infer, Apply Describe a ChIP-Seq...Ch. 18.7 - Prob. 1MICh. 18.7 - Retrieve, Infer, Apply Cite an infectious disease...Ch. 18.7 - Prob. 2CCCh. 18.7 - Prob. 3CCCh. 18 - Prob. 1RCCh. 18 - Prob. 2RCCh. 18 - Prob. 3RCCh. 18 - Prob. 4RCCh. 18 - Prob. 5RCCh. 18 - Prob. 1ALCh. 18 - Prob. 2ALCh. 18 - You are developing a new vaccine for a pathogen....Ch. 18 - Prob. 4AL
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- The amino acid asparagine is synthesized from aspartic acid by the enzyme asparagine synthetase (AS). In the previous problem you proposed a model for how this gene could be regulated. Suppose that you carry out an experiment to test your model. To do this you cut out the regulatory sequences upstream of the gene and fuse it to a gene for green fluorescent protein (GFP). Now you can visually observe when the gene is activated. You insert this engineered gene into a host cell and look for GFP expression. You discover some mutants that have different expression levels of GFP and call them GFP1- and GFP2-. The expression levels of GFP are given below. Cell GFP expression Wild type 100 GFP1- 50 GFP2- 0 Propose an explanation for these results based on your model. In other words, what was mutated and how? Your answer should include whether the mutation is (see links for more information): dominant or recessive https://www.ncbi.nlm.nih.gov/books/NBK21578/#A1877…arrow_forwardYou have isolated different mutants (reg1 and reg2) causing constitutive expression of the emu operon (which has genes emu1 and emu2). One mutant contains a defect in a DNA-binding site, and the other has a loss-of-function defect in the gene encoding a protein that binds to the site Say you don’t know which mutant has a defect in the site and which one has a mutation in the binding protein. To figure it out, you construct the two partial diploid strains (i and ii below), and you then assay the levels of the Emu1 and Emu2 proteins in these two strains. F’ (reg1- reg2+ emu1- emu2+) / reg1+ reg2+ emu1+ emu2- F’ (reg1+ reg2- emu1- emu2+) / reg1+ reg2+ emu1+ emu2- What proteins do you predict will be expressed for strains i and ii if reg2 encodes the regulatory protein and reg1 is the regulatory site?arrow_forwardIn lac operon, both gene A and gene B undergo a transcription process. Gene B can only undergo transcription in the presence of lactose and in the absence of glucose. The product of gene A is often altered by an inducer. Which of the following is true about genes A and B? Select one: a. Gene A= structural gene; Gene B= regulatory gene b. Gene A= regulatory gene; Gene B= structural gene c. Gene A= promoter gene; Gene B= operator gene d. Gene A= lacZ gene; Gene B= promoter genearrow_forward
- You have isolated different mutants (reg1 and reg2) causing constitutive expression of the emu operon (which has genes emu1 and emu2). One mutant contains a defect in a DNA-binding site, and the other has a loss-of-function defect in the gene encoding a protein that binds to the site. Is the DNA-binding protein a positive or negative regulator of gene expression?arrow_forwardYou are growing E. coli in a laboratory in order to study their operons. The growth media you are using contains lactose, no glucose and no tryptophan. Using your knowledge of operons and their regulation, answer the following questions Which operons would be functional under these conditions? 2. What repressors would be made? (NOTE: name the repressors using their gene names) 3.Which repressor(s) would be made in the inactive form? 4.Which repressor(s) would be made in the active form? 5.Which repressor(s) under these conditions can bind the operator sequence? 6.Which repressor(s)under these conditions cannot bind the operator sequence?arrow_forwardcAMP binds to cAMP Receptor Protein (CRP), allowing CRP to bind to the promoter of the lac operon a) in positive gene regulation by increasing the transcription when glucose is absent and lactose is present b) in negative gene regulation by decreasing the transcription when glucose is absent and lactose is present c) in positive gene regulation by increasing the transcription when glucose is present and lactose is absent d) in negative gene regulation by decreasing the transcription when glucose is present and lactose is absentarrow_forward
- Consider the trp operon found in E. coli. You have discovered a mutant strain of E.coli that contains a mutation in its trpR gene. This mutation prevents the trpR protein from binding to its small effector molecule. What would happen if both the wild type and mutant bacteria were put in an environment that contained lots of tryptophan? The normal (wild type) bacteria would have (i)________expression of the trp operon while the mutant bacteria would have (ii) _________ expression of the trp operon. A) (i) high (ii) high B) (i) low (ii) low C) (i) high (ii) low D) (i) low (ii) higharrow_forwardWhat would happen to the regulation of the tryptophan operon in bacterial cells that express a mutant form of the tryptophan repressor that (1) cannot bind to DNA, (2) cannot bind tryptophan, or (3) binds to DNA even in the absence of tryptophan?arrow_forwardThe lac operon has which of the following characteristics? O 1) usually requires an activator protein bound to the promoter to be transcribed. O 2) is inactive in the presence of lactose. O 3) is active in the presence of a repressor. O 4) Is a constitutively active operator. O5) is only active in the presence of lactose.arrow_forward
- Is the regulator protein that binds to the operator of this operon a repressor (negative control) or an activator (positive control)? Explain your reasoning.arrow_forwardYou can determine the regulation of an unknown operon based on descriptions like those in the table below. For each row, choose whether the fact would suggest positive or negative or inducible or repressible regulation and indicate your answer with an X in the column. Only one X should appear in each row. In the presence of the signal, the structural genes are expressed. In the presence of the signal, the structural genes are not expressed. In the absence of the signal, the structural genes are expressed. In the absence of the signal, the structural genes are not expressed. The binding of the regulatory protein to the operon results in structural gene expression. The binding of the regulatory protein to the operon prevents structural gene expression. A mutation in the regulatory protein results in constitutive expression of the structural genes. A mutation in the regulatory protein results in no or low expression of the structural genes. Positive? Negative? Inducible? Repressible?arrow_forwarda. What is the function of operons in bacterial gene regulation? b. Describe how a bacterial operon can be regulated by repressible proteins (such as the tryptophan operon). Include in your description both the “on” and “off” states of the operon. Key elements of your diagram should include: Co-repressor, Genes, mRNA, Operator, Operon, Promoter, Repressor, RNA polymerase.arrow_forward
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