Cell Biology (BIOL 3000) Exam II And All
Correct Answers.
How would you use epifluorescence microscopy to examine the localization of a chimeric GFP fusion
protein in a living cell? The λexcite for GFP is 395 nm & emission is 509 nm? - Answer Irradiate or
illuminate the cell at 395 nm and view it at 509 nm (Fig. 4-9)
Indirect immunofluorescence microscopy utilizes: - Answer A fluorescently tagged secondary antibody
that binds by its CDR to the Fc region of a primary antibody bound by its CDR to an epitope on its
corresponding antigen. (Fig. 4-14)
What proportion of the proteins encoded by the human genome are routed through the secretory
pathway? - Answer 33% (Part 2 Protein Trafficking Slide #1)
Where does "quality control" occur during the secretory pathway to eliminate misfolded proteins? -
Answer Within the ER (Part 2 Protein Trafficking Slide #1)
Refer to the figure attached below. Which of the following best describes the amino acid sequences that
comprise the N-terminal ER signal sequences found on most secretory proteins? - Answer They are
predominantly hydrophobic amino acids.
(https://collab.its.virginia.edu/access/content/attachment/720ac9b2-465f-4eb6-aba0-33d4cf1e5e74/
Tests%20_%20Quizzes/8168dc6f-0499-4741-8b34-04582ff3769a/ER%20Signal%20Sequences.pdf)
(ER signal sequences are comprised primarily of hydrophobic amino acids and the signal sequences are
cleaved from secretory proteins after they enter the ER.)
What distinguishes the membrane spanning α-helices found on the LDL receptor and Transferrin
receptor? - Answer The membrane spanning α-helix on the LDL receptor is a stop-transfer anchor. The
membrane spanning α-helix on the Transferrin receptor is an internal signal anchor.
(Part 2 Protein trafficking slides #15-17)
What does P54SRP indicate? - Answer It is the 54 kDa protein subunit of SRP.
, (Part 2 Protein Trafficking Slide #5)
How does a single P54SRP bind to diverse ER signal sequences that do not have identical amino acid
sequences? - Answer Multiple hydrophobic amino acids on P54SRP form van der Waals interactions
with "core" hydrophobic amino acids present on all ER signal sequences.
(Part 2 Protein Trafficking Slide #12)
What would you observe if the GTPs bound to both P54SRP and the α subunit of the SRP receptor were
replaced with GTP-γS? - Answer Translation of the nascent secretory protein would remain arrested
after the first 70 amino acids had been synthesized and the truncated protein would not enter the ER
through the translocon.
(Fig. 13-6, Protein trafficking slides #8-10)
The precursor to an IgG heavy chain accumulates in the cytosol of B lymphocytes if P54SRP cannot bind
to its ER signal sequence. Which pair of the following methods would definitely demonstrate if or how
the pre-IgG heavy chain that accumulates in the cytosol in cells where P54SRP cannot bind to the ER
signal sequence, differs from the heavy chain on a secreted IgG? - Answer SDS-PAGE + LC-MS/MS
(If P54SRP cannot bind to the ER signal sequence, the pre-IgG will be synthesized in its entirety and
accumulate in the cytosol and retain its N-terminal signal sequence. It will be ~1-2 kDa larger than
secreted IgG heavy chain. The size difference can be detected by SDS-PAGE (followed by a western blot
to visualize the proteins). The two proteins could also be distinguished by LC-MS/MS which would show
that the N-terminus of the pre-IgG begins with its ER signal sequence, but the N-terminus of the
secreted IgG begins with the first amino acid after the ER signal sequence was cleaved. For example, see
Protein trafficking slide #4. The N-terminus of mouse pre-IgG heavy chain is Met and the pre-IgG is 14
amino acids (~1400 Da) larger than secreted mouse IgG heavy chain which has an N-terminal Gly.)
Which of the following explains why cysteines located in the GTP binding site of the SRP receptor α
subunit have sulfhydral groups, while cysteines on the heavy and light chains of an antibody form
disulfide bonds? GSH = reduced glutathione; GSSG = oxidized glutathione. - Answer GSH:GSSG ratio in
cytosol is 50:1 & GSH:GSSG ratio in ER is 1:1
(Part 2 Protein trafficking slide #21)
You perform LC-MS/MS on the proteins present in a 100,000g X 60' pellet from lysed liver cells and
determined the following amino acid sequence for a hypothetical integral membrane protein. (The dots
merely denote every 10 amino acids beginning with M1 & ending with V120). Use this information to
Correct Answers.
How would you use epifluorescence microscopy to examine the localization of a chimeric GFP fusion
protein in a living cell? The λexcite for GFP is 395 nm & emission is 509 nm? - Answer Irradiate or
illuminate the cell at 395 nm and view it at 509 nm (Fig. 4-9)
Indirect immunofluorescence microscopy utilizes: - Answer A fluorescently tagged secondary antibody
that binds by its CDR to the Fc region of a primary antibody bound by its CDR to an epitope on its
corresponding antigen. (Fig. 4-14)
What proportion of the proteins encoded by the human genome are routed through the secretory
pathway? - Answer 33% (Part 2 Protein Trafficking Slide #1)
Where does "quality control" occur during the secretory pathway to eliminate misfolded proteins? -
Answer Within the ER (Part 2 Protein Trafficking Slide #1)
Refer to the figure attached below. Which of the following best describes the amino acid sequences that
comprise the N-terminal ER signal sequences found on most secretory proteins? - Answer They are
predominantly hydrophobic amino acids.
(https://collab.its.virginia.edu/access/content/attachment/720ac9b2-465f-4eb6-aba0-33d4cf1e5e74/
Tests%20_%20Quizzes/8168dc6f-0499-4741-8b34-04582ff3769a/ER%20Signal%20Sequences.pdf)
(ER signal sequences are comprised primarily of hydrophobic amino acids and the signal sequences are
cleaved from secretory proteins after they enter the ER.)
What distinguishes the membrane spanning α-helices found on the LDL receptor and Transferrin
receptor? - Answer The membrane spanning α-helix on the LDL receptor is a stop-transfer anchor. The
membrane spanning α-helix on the Transferrin receptor is an internal signal anchor.
(Part 2 Protein trafficking slides #15-17)
What does P54SRP indicate? - Answer It is the 54 kDa protein subunit of SRP.
, (Part 2 Protein Trafficking Slide #5)
How does a single P54SRP bind to diverse ER signal sequences that do not have identical amino acid
sequences? - Answer Multiple hydrophobic amino acids on P54SRP form van der Waals interactions
with "core" hydrophobic amino acids present on all ER signal sequences.
(Part 2 Protein Trafficking Slide #12)
What would you observe if the GTPs bound to both P54SRP and the α subunit of the SRP receptor were
replaced with GTP-γS? - Answer Translation of the nascent secretory protein would remain arrested
after the first 70 amino acids had been synthesized and the truncated protein would not enter the ER
through the translocon.
(Fig. 13-6, Protein trafficking slides #8-10)
The precursor to an IgG heavy chain accumulates in the cytosol of B lymphocytes if P54SRP cannot bind
to its ER signal sequence. Which pair of the following methods would definitely demonstrate if or how
the pre-IgG heavy chain that accumulates in the cytosol in cells where P54SRP cannot bind to the ER
signal sequence, differs from the heavy chain on a secreted IgG? - Answer SDS-PAGE + LC-MS/MS
(If P54SRP cannot bind to the ER signal sequence, the pre-IgG will be synthesized in its entirety and
accumulate in the cytosol and retain its N-terminal signal sequence. It will be ~1-2 kDa larger than
secreted IgG heavy chain. The size difference can be detected by SDS-PAGE (followed by a western blot
to visualize the proteins). The two proteins could also be distinguished by LC-MS/MS which would show
that the N-terminus of the pre-IgG begins with its ER signal sequence, but the N-terminus of the
secreted IgG begins with the first amino acid after the ER signal sequence was cleaved. For example, see
Protein trafficking slide #4. The N-terminus of mouse pre-IgG heavy chain is Met and the pre-IgG is 14
amino acids (~1400 Da) larger than secreted mouse IgG heavy chain which has an N-terminal Gly.)
Which of the following explains why cysteines located in the GTP binding site of the SRP receptor α
subunit have sulfhydral groups, while cysteines on the heavy and light chains of an antibody form
disulfide bonds? GSH = reduced glutathione; GSSG = oxidized glutathione. - Answer GSH:GSSG ratio in
cytosol is 50:1 & GSH:GSSG ratio in ER is 1:1
(Part 2 Protein trafficking slide #21)
You perform LC-MS/MS on the proteins present in a 100,000g X 60' pellet from lysed liver cells and
determined the following amino acid sequence for a hypothetical integral membrane protein. (The dots
merely denote every 10 amino acids beginning with M1 & ending with V120). Use this information to
