MOL 214

Exam 2
April 7, 2015

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Multiple Choice, 2 points each:

1. Below is shown a double-stranded segment of DNA. The transcription start site is

indicated by the nucleotides shown in the gray box. What are the first 10 nucleotides of the
transcript that is produced from this segment of DNA?

A. 5’-CAGUACAGUC-3’
B. 5’-CUGACAUGAC-3’
C. 5’-GUCAUGUCAG-3’
D. 5’-GACUGUACUG-3’

2. Which of the following occurs ONLY in prokaryotes?

A. An mRNA can be translated by more than one ribosome at one time

B. Transcription and translation are separated in space and time
C. Translation begins before transcription is completed
D. One gene can encode more than one polypeptide

3. You are studying a protein from a eukaryotic organism and are hoping to determine its
crystal structure. To do this, you need a large quantity of protein. You generate cDNA
encoding the protein, clone it into a plasmid, and introduce the plasmid into E. coli. When
you run an SDS-PAGE gel to visualize the protein, you find, to your disappointment, that the
protein was not produced by the bacteria. When you extract RNA from the bacterial cells
and do a Northern blot, you find that there is a large quantity of mRNA transcribed from
the recombinant plasmid. What could explain your findings?

A. Your plasmid does not have a promoter

B. Your plasmid does not have a ribosome binding site
C. Your plasmid contains a repressor site upstream of your gene
D. Your plasmid contains an activator site upstream of your gene

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4. Protein X will fold into its correct conformation only when protein Y is present.
However, protein Y is capable of folding into its correct conformation without protein X.
What is the function of protein Y with respect to protein X?

A. Chaperone
B. Ligand
C. Protease
D. Repressor

5. Why were temperature-sensitive mutants used in Schekman’s protein secretion

experiments?

A. He wanted to determine how temperature affects protein folding

B. Protein secretion is always dependent on the temperature
C. A blockage in protein trafficking and secretion leads to cell death
D. He used a mutagen that only generates temperature-sensitive mutations

6. Below is a graph showing the affinity of a receptor for three ligand molecules. Based on
this graph, which of the following choices is true?

A. Ligand “A” has a lower affinity for its receptor than Ligand “B”

B. More molecules of ligand “B” than ligand “C” would be required to reach the same
level of receptor activation

C. It would be more difficult to design a drug that prevented Ligand “C” from
binding to its receptor than a drug that prevented Ligand “A” from binding to its
receptor

D. The KD of Ligand “A” is lower than the KD of Ligand “C”

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7. Which of the following is TRUE, according to the operon model for gene regulation?

A. Genes coding for proteins that participate in the same process are coordinately
regulated
B. Each gene must have its own promoter in order to be transcribed
C. Genes encoding proteins that participate in different processes are often located
in the same operon
D. Each gene must have its own 5’ cap in order to be translated

8. Which of the following mutations would likely prevent Lac Repressor from binding to
the DNA?

A. A mutation in the promoter consensus sequence

B. A 5-base-pair insertion in the middle of the operator sequence
C. The deletion of one of the auxiliary operator sequences
D. A nonsense mutation in the repressor gene that results in only the DNA-binding
domain being translated

9. It is possible to create a pool of genomic DNA from cells by digesting the DNA into
fragments with a restriction enzyme. How will the the pool of genomic DNA differ from a
pool of cDNA generated from those cells?

A. the cDNAs will contain genes with both introns and exons
B. the genomic DNA will be more stable than the cDNA
C. the genomic DNAs will contain only the genes that are being actively expressed in
the cell it is made from
D. the cDNAs generated from this cell type may differ from the cDNAs generated
from another cell type

10. Which of the following questions can be answered by a DNA microarray experiment?

A. How many tyrosine molecules are in a particular protein

B. Which of two samples has greater abundance of a specific protein
C. Which of two samples contains genomic DNA
D. The number of mRNA molecules synthesized per second.
E. The relative abundance of specific mRNAs between two samples

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Short Answer:

11. For each of the following, provide one example of how the process differs in
prokaryotes and eukaryotes: (9 pts.)

(a) Transcription Initiation

(b) Transcription Termination

(c) mRNA Processing

12.

The DNA of a gene is shown above.

(a) Above, draw the longest mRNA that could be produced from this gene. Be sure to
label the parts of the mRNA, including the 5' untranslated region and the 3'
untranslated region. Under the mRNA, draw a line to indicate the protein coding
region (open reading frame). (5 pts.)

(b) Draw the shortest mRNA that could be produced. (1 pt.)

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13. There are 4 tRNAs for glycine. Would it be a problem for the cell if the aminoacyl tRNA
synthetase for one of the four glycine tRNAs was deleted? Why or why not? (3 pts.)

14. Explain how the results of Nirenberg and Leder’s experiment, which utilized single
codons, ribosomes, and radioactive amino acids to determine which codons coded for
which amino acids, would have been different if the genetic code did NOT possess the
characteristics listed below. Would their experiment have worked? Why or why not? (6
pts.)

(a) Degenerate

(b) Unambiguous

15. In 1-2 sentences explain…

(a) How prokaryotic ribosomes find the AUG on the mRNA. (3 pts.)

(b) How eukaryotic ribosomes find the AUG on the mRNA. (3 pts.)

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16. Clindamycin is an antibiotic that inhibits peptide bond formation. (6 pts.)

(a) Is clindamycin more likely to bind to the large or the small ribosomal subunit?
Explain your answer.

(b) Is clindamycin more likely to bind to ribosomal RNA or protein? Explain your
answer.

17. You are a graduate student studying transport of proteins by vesicles. You bathe a
pancreas tissue slice in a solution that contains radioactive leucine for 20 minutes and then
analyze the tissue by electron microscopy. Your electron micrographs show radioactive
protein in the rough ER and the Golgi. You report to your adviser that your protein was
transported from the ER to the Golgi. Why is she skeptical about your conclusion? What
would you have to do to convince her? (4 pts.)

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18. Below are three mutations in components of various signaling pathways. Would a
pathway that contained each of these mutations function? Why or why not? Be specific
about which step in the pathway would be affected, if any. (12 pts.)

(a) A mutant G Protein that can not hydrolyze GTP

(b) A mutant Receptor Tyrosine Kinase that can not form dimers

(c) A cell in which a response element for a hormone receptor has been deleted

19. In 2-3 sentences, describe 2 ways that expression of genes in prokaryotes can be
coordinately regulated. (4 pts.)

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20. You are investigating the metabolism of a novel sugar in your favorite bacterial species.
You have called this sugar Hershose after your favorite sugary treat, and you are interested
in understanding how the genes needed for Hershose utilization are regulated. Luckily,
you have a number of tools available to investigate this regulation. Below is shown the
upstream region of the DNA for the Hershose utilization operon. You have generated a
number of mutants in the upstream region in which various parts of the upstream region
have been deleted, shown below. The areas enclosed by the brackets ([ ]) represent the
region of the DNA that has been deleted.

You also have identified a compound (similar to X-Gal and called “X-Hersh”) that produces
a bright orange color when cleaved by the same enzyme that is responsible for
metabolizing Hershose. The colony colors that result from growth on plates containing the
X-Hersh compound and Hershose are shown below:

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In order to determine if any regulatory proteins bind to the upstream region of the
Hershose operon, you PCR this area from your mutant cells and perform a Gel-Shift Assay.
The results of this assay are shown below:

(a) Based on the colony color and gel shift data, where does the regulatory protein
most likely bind? Why? (3 pts.)

(b) Based on the colony color and gel shift data, what type of regulatory protein is
the Hershose regulator? Why? (3 pts.)

(c) What color would the colonies be if the regulatory protein for the Hershose
utilization genes was mutated so that it was always bound to the DNA? Why? (3
pts.)

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21. What is the purpose of incorporating a fluorescent label into the cDNA pool as part of a
microarray experiment? (4 pts.)

22. Briefly explain how DNA rearrangement and mRNA splicing account for antibody
diversity. (5 pts.)

23. Secreted proteins can be protected from degradation by proteases when they are
inside of vesicles. Propose an experiment that takes advantage of this property to show
that proteins translated on the rough ER end up in the lumen. Be sure to explain how you
will detect the protein. (6 pts.)

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