Transcript Bioluminescence - Fat Tuesday Productions

Bioluminescence
Uses and application in the real world
Uses
Bioluminescence Resonance Energy
Transfer (BRET)
 Bioluminescent molecules in gene
expression
 Identification of bacterial contamination
in food
 Artistic inspiration

BRET: Bioluminescence
Resonance Energy Transfer
BRET is a proximity-based
assay where the energy
generated by the catalytic
degradation of
coelenterazine by the
enzyme Renilla luciferase
(Rluc) (energy donor) is
transferred to a green
fluorescent protein (GFP)
acting as the energy
acceptor. The GFP then
emits light at its specific
emission wavelength.
Degradation of coelenterazine
by the enzyme Rluc
BRET
BRET: Bioluminescence
Resonance Energy Transfer

BRET can be used to observe protein-protein
interaction in living mammalian cells.
 It is based on the non-radioactive transfer of
energy between a luminescent donor Rluc
and a fluorescent acceptor (Green
Fluorescent Protein or GFP).
 May be used in the future to identify new
protein complexes in human.
Monitoring of ubiquitination in living
cells by BRET

Ubiquitination is descried as the rapid
process of post-translational modification
present in many aspects of biology involving
a covalent attachment of ubiquitin to proteins.
Monitoring of ubiquitination in
ling cells with BRET
Upon degradation of
DeepBlueC,the Rluc
fuses to β-arrestin and
emits blue light with an
emission peak at 395
nm. If the β-arrestin is
ubiquitinated, a transfer
of energy will occur
between Rluc and the
GFP2 fused to the N
terminus of ubiquitin,
resulting in re-emission
of light with an
emission peak at 510
nm.
Continuous light spectrum
Gene expression

Bioluminescence can be used to study
prokaryotic gene expression inside living cells
 It allows the observation of biological
processes in real time, as they happen.
 This technique can be used as a noninvasive
way to study protein trafficking, protein
function, genetic regulatory or image bacteria,
tumors and genes over a long period time.
Applications of bioluminescent imaging. (a) assessing the
levels of trans-gene expression, (b) the location and extent
of bacterial infection, (c) the efficiency of gene transfer and
expression, and (d) the trafficking patterns of lymphocytes.
Bioluminescence in gene expression

Luciferases are lightgenerating enzymes
that can be found in
bacteria, marine
crustaceans, fish and
insects.
 Luciferases are
nontoxic and can be
injected to become
gene expression
markers.
Gene Expression in Yeast Cells



Cells fused to GFP,
making the component
protein of microtubules
(alpha-tubulin)
The alpha-tubulin:GFP
fusion can be observed
by exciting the GFP,
causing a green light
emission.
It can be used to study
gene coding mutations
such as kinases.
Bioluminescence in Food
Industry
Methods of use

ATP
bioluminescence

Bacterial
bioluminescence
ATP Bioluminescence

Measures the amount of ATP that is
converted to photons of light by living cells.
 The amount of light emitted to proportional to
the number of bacteria in a food sample
 It can be used to measure the number of
lactic acid bacteria in a contaminated food
sample.
 Firefly luciferase is used to detect the
presence of ATP
Luciferin and the reaction with ATP
Bacterial Bioluminescence

Can be used to test for a specific bacterial
species or possible food born pathogen such
as Salmonella typhimurium.
 The lux gene, responsible for bacterial
bioluminescence, has been isolated and
cloned.
 The cloned lux gene can be injected into a
host-specific phage, which does not have the
ability to express the gene.
 If the phage infects a host bacterium, light
emission results.
Detection of Bacteria
Artistic Inspirations
Poetry
 Photography
 Painting

Fireflies, By Edgar Fawcett
I saw, one sultry night above a swamp,
The darkness throbbing with their golden pomp
And long my dazzled sight did they entrance
With the weird chaos of their dizzy dance!
Quicker than yellow leaves, when gales despoil,
Quivered the brilliance of their mute turmoil,
Within whose light was intricately blent
Perpetual rise, perpetual descent.
As though their scintillant flickerings had met
In the vague meshes of some airy net!
And now mysteriously I seemed to guess,
While watching their tumultuous loveliness,
What fervor of deep passion strangely thrives
In the warm richness of those tragic lives,
Whose wings can never tremble but they show
Those hearts of living fire that beat below!
Still Photography

Artistic re-creation of the
Bioluminescent Bay in Vieques
Sculpture
Montana State University-Bozeman
Bioglyphs project
 Collaboration between art and science
during 2002 by members of the center
for Biofilm Engineering and the MSU
School of art

Lights on
Lights off
Lights on
Lights off
Bioglyphs

Involves the practice of "painting" on
prepared Petri dishes with a sort of "invisible
ink" composed of liquid medium inoculated
with the bacteria.
 The microorganisms themselves went to
work, multiplying on the plates and beginning
to produce light within 24 hours.
 The only light available to view the art was
that produced by the bacteria themselves.
Over the five-day period, the light intensity of
the paintings changed as the bacteria
multiplied and then gradually consumed the
nutrient available.
Gallery, with lights on
Gallery, with lights off
Sources

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Stephane Angers, Ali Salahpour, Eric Joly, Sandrine Hilairet,
Dan Chelsky, Michael Dennis, and Michel Bouvier. “Detection
of β 2- adrenergic receptor dimerization in living cells using
bioluminescence resonance energy transfer (BRET).”
Proceedings of the National Academy of Sciences 97(2000):
3684-3689. http://www.pnas.org/cgi/content/full/97/7/3684 (5
October 2007)
Claire Normand1, Stéphane Parent1, Benoit Houle, Anne
Labonté, Lucie Bertrand, Mireille Caron, Mireille Legault,
Stéphane Angers, Michel Bouvier, Erik C. Joly and Luc
Ménard. “BRET2™: Bioluminescence
Resonance
Energy Transfer, a Novel Assay Technology to Examine
GProtein Coupled Receptor Activation in Intact Cells.” 2002
http://las.perkinelmer.com/Content/RelatedMaterials/Posters/
PSH_BRET2NovelAssayTechnology.pdf (9 October 2007).
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Saman Hosseinkhani. “Bioluminescnece.” 2007
http://www.modares.ac.ir/sci/saman_h/Pages/Bioluminescence
%20.htm (7 October 2007).
M. W. Griffiths. “Applications of Bioluminescence in the dairy
Industry.” Journal of Dairy Science 76 (1993): 3118-3125.
http://jds.fass.org/cgi/reprint/76/10/3118.pdf (2 October 2007).
Julie Perroy, Stephanie Pontier, Pascale G Charest, Muriel
Aubry and Michel Bouvier. “Real-time monitoring of
ubiquitination in living cells by BRET.” 1 (2004): 203-208.
http://www.nature.com/nmeth/journal/v1/n3/abs/nmeth722.html;jsessio
nid=A04B9606CE4D3B8FB5147E17BDEF094D (6 October 2007).
Bouvier;s Lab. “Using BRET to study Protein-Protein interactions.”
June 5, 2005.
http://www.mapageweb.umontreal.ca/bouvier/bret/index.html (8
October 2007).
Montana State University. “Bioglyphs.” January 3, 2007.
http://www.erc.montana.edu/Bioglyphs/default.htm (9 October 2007).
Jackie Vogel. “Research.” June 3, 2005.
http://biology.mcgill.ca/faculty/vogel/research.html (10 October
2007)