3.4 study guide Flashcards
You are studying a fluorescent protein. Which must be true about your protein?
a. It is excited by light energy
b. It is excited by mechanical energy
c. It is excited by chemical energy
d. It must bind calcium to fluoresce
e. All of the above
It is excited by light energy
A fluorescent protein emits light in the green wavelength range. In which of these possible color ranges
could its excitation wavelength be?
a. Red
b. Yellow
c. Blue
d. Any of these
e. Fluorescent proteins are not excited by light
Blue
What is the chromophore of aequorin?
a. 11-cis retinal
b. The amino acids S65, Y66 and G67
c. Heme
d. Coelenterazine
e. Calcium
Coelenterazine
Which is FALSE about the GFP?
a. The fluorophore is composed of the amino acids S65, Y66 and G67
b. It is a beta barrel protein
c. It absorbs a green photon
d. The fluorophore forms an N-imidazolinone ring
e. The fluorophore is inside the barrel
It absorbs a green photon
The mature GFP chromophore involves:
a. An acyclimine ring
b. An imidazolinone ring
c. Coelenterazine
d. No cyclic ring system
e. None of the above
An imidazolinone ring
Your lab has a GFP reporter gene, that you decide to convert to BFP by mutating residue 66 to histidine.
Why does this mutation affect the color of fluorescence?
a. It creates a fluorophore that emits more energy after it is excited
b. It creates a fluorophore that emits less energy after it is excited
c. It exposes the fluorophore to the aqueous environment of the cell
d. It causes the fluorophore to be excited by chemical energy
e. All of the above
It creates a fluorophore that emits more energy after it is excited
Your professor asks you to fuse the protein you’re studying to an RFP so that you can visualize it in the
cell. Your lab has both mCherry and DsRed, and you have to decide based on the properties of these two
fluorescent proteins which one is best for this application. Which do you decide to use for your fusion
protein and why?
a. mCherry, because it is less prone to aggregation
b. DsRed, because it is less prone to aggregation
c. DsRed, because it is faster to fold
d. mCherry, because it is a tetramer
e. Either one is OK, both DsRed and mCherry would work very well as a fusion protein
mCherry, because it is less prone to aggregation
You are making mutations in mCherry to study the effects of these mutations on protein structure and
function. You find that one of the mutations you make creates a large gap in the barrel structure, leaving
the interior exposed to the aqueous environment of the cell. What effect on fluorescence would you
expect this mutation to have?
a. The fluorescence emission wavelength will shift over time
b. The fluorescence excitation and emission wavelengths would be different
c. The fluorescence emission will be brighter
d. The fluorescence emission will be quenched
e. None of the above
The fluorescence emission will be quenched
You’re studying a protein and want to find out whether it is stable for long periods in the cell after it is
translated. Which fluorescent-based protein sensor would you use?
a. A sensor with a large gap in the barrel
b. A sensor that has a fluorophore that changes its color of emission over time
c. A sensor that has a small hole in the barrel
d. A sensor that is sensitive to changes in pH
e. Any of these would work well
A sensor that has a fluorophore that changes its color of emission over time
You would like to design a FRET sensor system. If you use green fluorescent protein as your photon
donor, which of these proteins is least likely to be useful as a photon receiver?
a. Yellow fluorescent protein
b. mOrange (orange)
c. mCherry (red)
d. Blue fluorescent protein
e. Any of these would work equally well
Blue fluorescent protein
You are studying a signaling pathway that produces cAMP and would like to know whether cAMP-
dependent protein kinase A (PKA) is activated. To do this you decide to use a FRET sensor system: you
fuse CFP to the PKA regulatory domain and YFP to the PKA catalytic domain. If you see cyan
fluorescence, but not yellow fluorescence after you excite CFP, what conclusion can you draw from your
experiment?
a. PKA is activated
b. PKA is not activated
c. cAMP is not made
d. The fusion protein you made failed
e. None of the above
PKA is activated
You would like to measure changes in calcium levels inside a cell. To visualize calcium levels, you
decide to use the monomolecular FRET sensor Cameleon. Which protein has CFP and YFP fused to
either end to make the Cameleon sensor?
a. PKA
b. Calmodulin
c. Calreticulin
d. A calcium-specific ion transporter
e. SERCA
Calmodulin
You are designing a FRET sensor to examine the activity of the MAP kinase ERK. Which of these
domains does your sensor need to have?
a. Fluorescent proteins fused to either end
b. An ERK substrate domain
c. A phospho-binding domain
d. A flexible linker domain
e. All of the above
All of the above
