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03/21/2015
CONTENTS
HOT SPRINGS OF
YELLOWSTONE NATIONAL PARK
Section
Topic
1………………………………….. The Archaea
ARCHAEA
2………………………………….. Extremophiles
3………………………………….. Significance to Astrobiology
4……………………………….... Applications and Current Research
Jared Allyn
^
DISCOVERY
Result:
Archaebacteria
+
Eubacteria
FALSE-COLOR TRANSMISSION
ELECTRON MICROGRAPH IMAGE
CHARACTERISTICS OF AN UNKOWN MICROBE
Bacteria
Unknown Group of
Microbes
Eukarya
Nuclear Envelope
No
No
Yes
Organelles
No
No
Yes
Cell Wall (With
Peptidoglycan)
Yes (Yes)
Membrane Lipids
Unbranched Hydrocarbons
(Ether Linked)
Some Branched
Hydrocarbons
(Ester Linked)
Yes (No) –
Unbranched Hydrocarbons
(Ester Linked)
No
RNA Polymerase
One Type
Many Types
Many Types
Amino Acid for Protein
Synthesis Initiation
Formyl-methionine
Methionine
Methionine
In Some Genes
Yes
The Cell Wall
• Cell walls are outermost layers
Pseudopeptidoglycan
Introns
No
Response to Streptomycin
& Chloramphenicol
Growth Inhibited
Growth Not Inhibited Growth Not Inhibited
Histones w/DNA
No
Yes
Yes
Circular Chromosome
Yes
Yes
No
• Protection and stability
• Found in almost all
microorganisms and
some Eukaryotes, like
plants
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The Cell Wall
The Cell Wall
Bacteria
Unknown Group of Microorganisms
• Nearly all bacteria
contain a cell wall
*One common exception is genus Mycoplasma
e.g. Mycoplasma pneumoniae
• Cell wall
composition
varies
• All cell walls contain
peptidoglycan
• All cell walls lack
peptidoglycan
• Separated into Gram+
and Gram-
• Composition of
cell wall provides
support from
external stresses
A DOMAIN OF ITS OWN
A DOMAIN OF ITS OWN
The old classification method
used biochemical, metabolic,
& morphological distinctions
to classify organisms
Modern Approach:
Phylogenetics
Genetic comparison through
rRNA sequencing
Genetic differences were
too great to accurately
include Archaebacteria
and Eubacteria in the
same Kingdom, but similar
enough to be considered
a Prokaryote like bacteria
Eubacteria
Archaebacteria
- Biochemical Makeup
e.g. Cell Walls
Result: Fail to Distinguish from Bacteria
Figure 5.11
Page 168
- Metabolism
e.g. Energy Consumption
Result: All Fall Under a
Metabolic classification of Life
- Morphology
e.g. Cell Shape
Result: Fail to Distinguish from
Bacteria
…thus, a new
Domain/Kingdom was born.
Figure 5.11
Page 168
ENVIRONMENTS
BLACK SMOKER
Thermophiles
(Deep-sea Hydrothermal Vent)
Require temperatures between 40-700C
(104-1580F)
*Heat sterilization occurs at ~160-1800F
Acidophiles
Optimal growth at low pH (~2 or
below)
*Black coffee (~pH 5) --> Orange
Juice (~3.5) --> Soda (~2.5) -->
Battery Acid (~1) --> 1N Sulfuric
Acid (~0.3)
Alkaliphiles
Halophiles
Optimal Growth at high pH, alkaline, conditions
(~8.5-11)
*Baking Soda Solution (~8.4) --> Detergent (~10.8)
--> Clorox (~12.5)
Extreme halophiles can
grow at 3.4-5.1M (or
20-30% concentration)
aqueous NaCl.
Seawater is 0.6M NaCl
(3.5% concentration)
*Archaea are primarily extremophiles, a group
that contains some bacteria and eukarya
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SURVIVING EXTREME CONDITIONS
SURVIVING EXTREME CONDITIONS
Acidophiles
THERMOPHILES
H+ Pump Efficiency
TAQ Polymerase
SURVIVING EXTREME CONDITIONS
SURVIVING EXTREME CONDITIONS
Halophiles
Alkaliphiles
Extreme Osmoregulation
H+ Favoring Na/H+ Antiporters
(Active Acidification)
*Passive acidification from acidic components of cell wall
OTHER ARCHAEA
Some archaea thrive in moderate conditions
(20-450C or 68-1130F) such as the environments
human beings occupy. These species are known
as “mesophiles”
Other archaea thrive in extreme cold (-20-100C)
and are known as “psychrophiles”
THE YOUNG EARTH
An Inhospitable World
Early Bombardment Period
(4.1-3.8 bya): Hydrothermal vents
were numerous and extremely
active
The first organisms to occupy the
planet were unicellular
chemolithoautotrophs
~3.5 bya: LUCA splits off into
Archaea and Bacteria
Hyperthermophiles and
methanogens provide insight
DEEP-SEA HYDROTHERMAL VENT
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THE HYPERTHERMOPHILE
Strain 121
- Thrive at temperatures ~800C and above
Section 3
SIGNIFICANCE TO ASTROBIOLOGY
“The combined implications of their
[archaea] evolutionary role and their capacity
to thrive in enviromental extremes have
expanded the horizons for the astrobiology
community in searching for extraterrestrial
life.” – Ricardo Cavicchioli from Archaea:
Molecular and Cellular Biology
VOYAGER
- Many not only survive, but successfully
reproduce at temperatures 1000C and
above
- Some are found in colonies on the
superheated walls of hydrothermal
vents
- These organisms are often found at
great depths within the ocean where
pressure prevents water from boiling
SIGNIFICANCE TO BIOLOGY
• Large contribution to C, N,
and S cycling
• Changed the way
microbiologists see life
– Methanogens a major source
of atmospheric methane
– Phylogenetics
– 3-Domain system
– Ubiquity
• Relatively unknown
providing an abundance of
research potential
• Unique relationships with
other organisms
– No known pathogenic archaea
– Mutualism: interaction
between methanogenic
archaea and protozoa in
termites
Section 4: APPLICATIONS & CURRENT RESEARCH
Significance to Biology
Importance to the
Search for
Extraterrestrial Life
EUROPA
IMPORTANCE TO THE SEARCH FOR
EXTRATERRESTRIAL LIFE
• The archaea have expanded the boundaries of life’s
survivability
• They have allowed for the consideration of more
inhospitable habitats
• Have provided new insights into the formation of organisms
opening up the possibilities for the universality of life
Some of the habitats where archaea and other extremophiles
have been found are not too dissimilar from those located
on Mars and what is theorized to exist within Europa.
CURRENT RESEARCH
Here, at SDSU…
Dr. Richard Bizzoco
Professor of Biology
Specializes in the classification of new
organisms in extreme environments
• Current Research
– Richard Bizzoco
Professor of Biology, Ph.D.
DNA
• Specializes in the classification of new
organisms in extreme environments
– Jared Allyn, Volunteer
• Challenges with imaging
TAQ Polymerase
• TAQ Polymerase and more
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JARED ALLYN
JARED ALLYN
WHAT DO I DO?
Volunteer Researcher in Microscopy
Transmission Electron Microscopy
Volunteer Researcher in Microscopy
…to produce images like this.
I operate one of these…
Blood Vessel from
mouse liver tissue
What do I do?
Why is my work beneficial?
Goals In Relation To Astrobiology
WHAT DO I DO?
Pellet
T4 Bacteriophage
(negative stain)
Transmission Electron Microcope
WHAT DO I DO?
Centrifugation creates a pellet of material including microbial cells
- Animals or plant tissue must be
extracted and diced
- Microbes must be sedimented
- Cells must be preserved or
fixated to appear “natural”
-
Chemical to cross-link molecules
within cell
Causes long-term preservation
and image contrast
Continued
- Cells must be washed with
water to remove fixatives
- Cells must then be washed in
alcohol to remove water
- Cells go through a process to
embed them in plastic
maintaining cell integrity
- Ultra-thin plastic sections
trimmed in a “microtome”
~1mm3 tissue sample
Finally, the sample
is ready to view
under the
microscope
Effects of different fixation
procedures on contrast and image
clarity
Challenges with Imaging Extremophiles
• Since extremophiles thrive in inhospitable
environments, they are very difficult to
prepare for the electron microscope
• Many of the organisms I image are acidophilic
thermophiles
– Biological fixatives are converted to acids at high
pH
– Fixatives cross-link too quickly at high
temperatures
WHY IS MY WORK BENEFICIAL?
Challenges with Imaging Extremophiles
• I am learning how to image these organisms in
order to find ways to improve image quality
• This requires taking detailed notes on how I
prepare specimens in order to make necessary
adjustments to protocol
• I also provide Dr. Bizzoco a helping hand in
imaging his specimens, which is a process that
takes days
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Applications
CONTRIBUTING TO ASTROBIOLOGY
• Electron micrographs provide important data
• This data can be used to find new biological
structures and help describe biochemical
processes
• These structures can provide insight into how
organisms withstand such inhospitable
environments
Applications
Biotech
• Extremozymes
– Ex. Hyperthermophilic proteases increase the cleaning
ability of detergents at high temperatures
• Archaeosome Vaccines
- Using highly stable archaeal membrane to deliver drugs and
vitalize immune response
• Bioremediation
- Treating contaminated soil and waste water
• Play a role in anaerobic decomposition
TAQ Polymerase
• DNA polymerases
vital to DNA replication
• Found in Thermus
Aquaticus (“TAQ”)
DNA
950C
Denaturation
600C
600C
Annealing
Annealing
– Thermostable
• Polymerase Chain Reaction
(PCR)
-
Generates numerous copies of a
single sequence
Uses: diagnosing hereditary
diseases, phylogeny, IDing genetic
fingerprints, and more
720C
Elongation
TAQ Polymerase
Repeat
WHAT TO REMEMBER
• Archaea are prokaryotes along with bacteria, but
constitute a 3rd domain of life
• This is due to many distinctions from bacteria that
apply to almost all archaea
• The archaea have mostly been found in extreme
environments that are either high in acidity, basicity,
temperature, or salt concentration
– Sewage treatment facilities
• S-layer glycoproteins
-Self-assembly could benefit nanotechnology
-Important filtration capacity due to exclusion properties
• These organisms expand the limits of life’s capability to
survive and thrive making them extremely important to
the search for extraterrestrial life.
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