Exam Code Number:____________

MOL 214
EXAM 2
April 7, 2011

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Multiple choice questions

PLEASE NOTE ONLY ONE CHOICE IS CORRECT (10 points)

1. A protein is synthesized from its _______ as the ribosome moves toward the ______ end of
the mRNA.
a. N to C terminus; 5’
b. C to N terminus; 5’
c. N to C terminus; 3’
d. C to N terminus; 3’
e. none of the above

2. How is initiator tRNA unique among tRNAs?

a. It is the only tRNA that never sits in the A site of the ribosome.
b. It is the only tRNA that never sits the P site of the ribosome.
c. It is the only tRNA with arginine.
d. It is the only tRNA without an amino acid attached.
e. It is the only tRNA with serine attached.

3. DNA that is synthesized using mature mRNA molecules as template is called

a. rDNA
b. gDNA
c. cDNA
d. mDNA
e. hDNA

4. The RNA-induced silencing complex (RISC) with miRNA or siRNA affects gene expression
by binding to
a. mRNA
b. rRNA
c. snRNA
d. tRNA
e. protein product translated from the mRNA

5. Scientists discovered a mutation that causes the tRNA molecule that recognizes cysteine
codons (tRNA.Cys) to be mischarged such that it actually carries alanine (Ala-tRNA.Cys). What
would happen to proteins synthesis in cells with this mutation?

a. The protein will contain Cys in place of Ala.

b. The protein will contain Ala in place of Cys.
c. Protein synthesis would stop.
d. Cys will be incorporated randomly at any codon.
e. Ala will be chemically converted to cysteine by cellular enzymes.

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Short answer questions (Gavis Lectures)

1. Explain how hydrogen bonding is used to create an alpha helix structure in a polypeptide
chain.(2 points)

2. Mutations that prevent phosphorylation of RNA polymerase CTD are lethal to mammalian
cells. Give two specific reasons why these mutations are so deleterious. (4 points)

3. What are two ways in which ribosomes and spliceosomes are similar to each other?(2 points)

4. Eukaryotic mRNAs are often shorter than the DNA sequence from which they are
transcribed. Why is this? (2 points)

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5. The following questions pertain to the lac operon in E.coli.(10 points)

(a) The lac O operator sequence overlaps a portion of the lac operon promoter. Why is this
configuration important for the regulation of the lac operon?

(b) How does an inducer work to turn on transcription of a bacterial operon that has been
repressed, such as the Lac operon?

(c) When E. coli cells are growing in the presence of lactose with very little glucose, is the level
of cAMP high or low? Circle one

High Basal None

(d) What is the level of lac operon transcription in presence of high glucose and low lactose
when there is a mutation in the operator sequence that cannot bind Lac repressor. Circle one

High Basal None

(e) What is the level of lac operon transcription in presence of low glucose and high lactose
when the promoter sequence is mutated so that it cannot bind RNA polymerase? Circle one.

High Basal None

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6. What effect would the following changes have on the expression of a gene?

a) if the promoter sequence is inverted? Explain your answer. (2 points)

b) if its enhancer sequence is inverted? Explain your answer. (2 points)

7. Give two reasons why chemicals that inhibit GTP hydrolysis prevent translation. Be sure to
indicate the step in the translation process that is affected. (4 points)

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8. Your thesis advisor discovered a mutation in a gene called Bafld that causes short term
memory loss in mice. You want to know more about this gene because you think that the
human version might be involved in retention of facts you memorize for exams. You isolate
a large (6 kb) restriction fragment of DNA that you believe contains the normal (non-mutated
version) gene from a mouse.

a) A lab mate has isolated mRNA from 7 different tissues in the mouse. Briefly describe an
experiment you could perform to verify which tissue or tissues express this gene? (3 points)

b) Your experiment shows that this gene is actually expressed in three different tissues. You
want to determine whether there are different enhancers responsible for this pattern of
expression. Explain how you would go about identifying such enhancers experimentally. (4
points)

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c) Your advisor has done some experiments that indicate that the Bafld protein is a transcription
factor. She has also done a microarray experiment, comparing RNA extracted from tissue from
normal mice (these samples are labeled in green) with RNA extracted from tissue from mice that
are mutant for Balfd (these samples are labeled in red).

You look at the data from the microarray. Most of the spots are yellow, but you find 10 spots
that are green and 1 spot that is red.

i. What do the green spots signify? (2 points)

ii. What does the red spot signify? (2 points)

iii. Based on what you know about the Bafld protein, what is a reasonable explanation for
the 10 green spots in the microarray experiment? (2 points)

9. Arginine has 6 codons: AGA, AGG, CGU, CGC, CGA, CGG. (4pts)
a. What is the minimal number of tRNAs needed to decode Arg?

b. What are the anti-codon sequences of those tRNAs? Be sure to indicate the 5’
and 3’ ends of each anticodon.

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How we study cells (10 points)

10. Imaging is a major tool in studying organelles and molecules in cells. Advances in imaging
technology have developed tools with greater resolution.
a. The maximum resolution of a light microscope is approximately 0.3µm. What are 2 of
the parametersthat determine this resolution? (2 points)

b. A specimen can be viewed dry, under water immersion or under oil immersion in light
microscopy. Which gives a better resolution? Air, water and oil have refractive indices
of 1.00, 1.33 and 1.51, respectively. (1 point)

c. What does θ represent and what are two things that can be done to improve θ to increase
resolution? (3 points)

d. If mammalian cells in culture have been transfected with the gene for the Green
Fluorescent protein (GFP), what optical technique would you use to see the expression of
this protein? (1 point)

11. The electron microscope is an instrumental tool in analyzing small structures.

a. Why does the electron microscope have far better resolution than the light microscope?
(1 point)

b. How does the electron microscope focus the image? (1 point)

c. What is the typical size of the smallest structures that can be visualized by EM? (1 point)

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Cell Cycle and Cancer (13 points)

12. Experiments using a cell free system in Xenopus oocytes from Kirshner and colleagues,
combined with Hunt’s experiments in sea urchin eggs, partially purified an activity that is
crucial in determining whether cells undergo mitosis.
e. What were the two proteins in this activity? (2 points)

f. What are the levels of these two proteins at the start of mitosis? (1 point)

g. How do the amounts and activities of these proteins change after a cell has gone
through mitosis? (2 points)

13. You are a scientist trying to identify cell division cycle regulatory genes. You approach it
using classical genetic methods to generate temperature-sensitive mutants. You identify a
yeast cell carrying a temperature-sensitive mutation in a gene required for stabilization of the
microtubules.

h. If cells were incubated at the restrictive temperature, at what stage of the cell division
cycle would you expect the mutant cells to accumulate? Briefly explain your answer
(2 points)

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i. The diagram below demonstrates the temperature-sensitive mutant incubated at

permissive (25oC) and their arrest phenotype at the restrictive temperature (36oC).
The cells arrested as either one or two large-budded cells.
Place an arrow representing the execution point for this mutant. (1 point)

c) What is the explanation for the different ways that the cells arrested? (2 points)

14. The tumor suppressor protein, retinoblastoma (RB), holds cells in G1 phase until it gets a signal for
cell proliferation.
j. What is the mechanism of RB inhibition on the cell cycle? (2 points)

k. Why does having a mutation in only one copy of RB not result in cancer? (1 point)

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Secretory pathway (10 points)

15. You are a scientist working on the protein secretory pathway and decide to make
temperature-sensitive yeast mutants to examine this system. Invertase is a normally secreted
protein, and it gets glycosylated/modified along the secretory pathway in ways that affect
how it runs on a protein gel.
You identified two mutants, A and B, which block secretion of invertase. Results from the
western blot analysis of invertase from each mutant at permissive and restrictive
temperatures are shown below.

a) Predict where in the secretory pathway is Mutant A blocked. Based on what you learned in
lecture, which protein might be defective in this mutant? (2 points)

b) Predict where in the secretory pathway is Mutant B blocked. Based on what you learned in
lecture, which protein might be defective in this mutant? (2 points)

c) How would you confirm the order of invertase secretion using the two mutant strains? (2
points)

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16. You are a scientist working on the protein secretory pathway and you decide to make
temperature-sensitive yeast mutants to examine this system.
a) After mutagenizing yeast strains and looking for no growth at the restrictive temperature,
how could you identify cells that are mutant in protein sorting? (2 points)

b) After much hard work, you have identified a temperature-sensitive mutant that does not
grow at the restrictive temperature and has proteins accumulating in the ER. From what you
have learned about how proteins are made and secreted to other organelles, suggest a protein
and its mechanism that might be defective in this mutant. (2 points)

Cytoskeleton (12 points)

17. State the 3 major classes of filaments that make up the cytoskeleton and state the main
protein subunit that each filament is composed of. (3 points)

b) Which cytoskeletal filaments would be most plentiful in a muscle cell? (1 point)

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18. Dynamic instability causes microtubules to either grow or to shrink rapidly. Consider an
individual microtubule that is in its shrinking phase.
What must happen at the end of the microtubule in order for it to stop shrinking and start
growing? (2 points)

19. The formation of actin filaments in the cytosol is controlled by actin-binding proteins. Some
actin-binding proteins significantly increase the rate at which formation of actin filaments is
initiated.
a) Name a protein that initiates such a filamentous growth (1 point).

b) Briefly describe its mechanism in forward movement of a lamellipodium (2 points)

20. You work in a lab studying motor proteins and you think you have just discovered a new
microtubule motor protein. You have purified this protein, and you borrow from a lab mate
axonemes purified from flagella and fluorescently labeled tubulin.

a) How could you determine in vitro which end of the axoneme is the plus end? (1 pt)

b) How would you determine whether this newly discovered microtubule motor moves
preferentially towards the plus or minus end? (2 points)

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