MCAT Topic Focus Biology and Biochemistry

Biology: Electrophoresis and Blotting Passage 1 2 3 4

(Questions 1-6)
TOP
Vesicular Stomatitis Virus (VSV) is a costly agri­ cultural
pathogen that infects farm animals such as pigs, sheep, and
cattle, causing a flu-like illness. When VSV enters a eukaryotic
cell, the virus takes over the protein synthesis machinery of the
host cell to make its own pro­teins. The VSV genome consists
of five genes. Only one of these encodes an integral membrane
protein, found in the plasmalemmas of infected cells, the VSV-G BOTTOM
protein. This is a structural protein with no enzymatic activity.
It has a large extracellular domain, which is glycosylated, a
small transmembrane region, and a very short cytoplasmic tail. Figure 1 Results of Gel Electrophoresis Experiment
Eukaryotic cells made to express VSV-G alone and no other VSV
proteins do not exhibit any change other than the expression of Adapted from Microbiology, by L. M. Prescott, J. P. Hartley, and D. A. Klein.
©1990 Wm. C. Brown Publishers. pp. 118-120.
VSV-G.

VSV was used in four in vitro translation experiments to

study how eukaryotic integral membrane proteins are trans­
1. Which of the following is the best justification for the
lated and processed. VSV-G mRNA and all eukary­otic com­
ponents necessary for translation were placed in a medium
with a composition similar to that of host cell cyto­plasm. The
eukaryotic components were isolated from uninfected eukaryotic
cells. Rough endoplasmic reticulum (RER) was left out of some
experiments but included in others.
In Experiment 1, VSV-G mRNA was combined with all the
eukaryotic translation components except RER; the polypeptide
produced in this experiment was denoted G0.
In Experiment 2, RER was included, and the product was
denoted G2.
In Experiment 3, the G2 protein was synthesized once again,
but in the presence of trypsin, in order to determine whether any
of the protein domains were protected by the RER. Trypsin is a
proteolytic enzyme (hydrolyzes poly­peptides), that cannot cross
the RER. The trypsin was removed before the G2 protein was
separated from the RER.
In Experiment 4, G0 is synthesized (RER not added), in the
presence of trypsin.
Figure 1 below shows the results of gel electrophoresis of the
polypeptide products of the experi­ments. The larger the protein,
the more slowly it runs from the top of the gel to the bottom. The
columns, or lanes, of the gel are labeled according to experiment
number.
use of VSV-G in attempts to understand the translation of
eukaryotic integral membrane proteins?
A. Viral ribosomes closely resemble eukaryotic
ribosomes.
B. VSV-G can be translated in vitro.
C. Viruses possess no protein synthesis machinery of
their own, and must borrow it from the host.
D. VSV-G is an important structure in the eukaryotic
cell membrane.
2. Which of the following is the best explanation for the
difference in apparent molecular weight of VSV-G protein
produced in Experiments 1 and 2?
A. The hydrophobic transmembrane domain of VSV-G
is removed by a protease in Experiment 2.
B. A hydrophobic signal peptide for entry into the RER
is removed by a protease in Experiment 2.
C. VSV-G was reduced in size by treatment with
trypsin.
D. VSV-G was glycosylated in Experiment 2 but not in
Experiment 1.
3. What is the most likely explanation for lane 4 being
empty in Figure 1?
A. No protein was synthesized in Experiment 4.
B. The polypeptide produced in Experiment 4 is
present, but not seen.
C. The product of Experiment 4 was not loaded onto
the gel.
D. The product of Experiment 4 was degraded by
trypsin.

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 MCAT Topic Focus Biology and Biochemistry

4. Which domain of the VSV-G protein would be most 6. Based on the results of the experiments described in
accessible to host antibodies? the passage, which of the following provides the best
support for the hypothesis that the mechanism of in vitro
A. Cytoplasmic domain translation had not been altered by VSV proteins?
B. Transmembrane domain
C. Glycosylated domain A. All eukaryotic cellular components came from
D. Enzymatic domain uninfected cells, only VSV-G mRNA was added to
the in vitro translation mixture, and the function of
the VSV-G protein was known not to affect cellular
5. Which of the following is the best analysis of the results? components.
B. All eukaryotic cellular components came from
A. G2 is smaller than G0 due to the absence of RER in infected cells, only VSV-G mRNA was added to
some of the experiments. the in vitro translation mixture, and the function of
B. G0 is smaller than G2 due to the absence of RER in the VSV-G protein was known not to affect cellular
some of the experiments. components.
C. G2 is smaller than G0 due to the absence of VSV-G C. All eukaryotic cellular components came from
mRNA in some of the experiments. uninfected cells, all VSV mRNAs were added to
D. G0 is smaller than G2 due to the absence of VSV-G the in vitro translation mixture, and the function of
mRNA in some of the experiments. the VSV-G protein was known not to affect cellular
components.
D. All eukaryotic cellular components came from
uninfected cells, only VSV-G mRNA was added
to the in vitro translation mixture, and the function
of the VSV-G protein was known to affect cellular
components.

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