extremophiles - rtb 11.3
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Introduction to Extremophiles
• What are Extremophiles – Live where nothing else can
• How do they survive?– Extremozymes (more details later)
• Why are they are interesting?• Extremes fascinate us– Life on other planets– Life at boiling temperatures
• Practical applications are interesting
– Interdisciplinary lessons• Genetic Prospecting
Extremophile
• Definition - Lover of extremities• History–First suspected in 1950’s–Extensively studied since
1970’s• Temperature extremes–Boiling or freezing, 1000C to -
10C –Chemical extremes –Vinegar or ammonia (<5 pH or
>9 pH)–Highly saline, up to x10 sea
water• How we sterilize & preserve
foods today
Extreme Temperatures
• Thermophiles - High temperature– Thermal vents and hot springs–May go hand in hand with chemical
extremes
• Psychrophiles - Low temperature– Arctic and Antarctic
• 1/2 of earth’s surface is oceans between 1-40C
• Deep sea –10C to 40C• Most rely on photosynthesis
Chemical Extremes
• Acidophiles - Acidic– Again some thermal vents & hot springs
• Alkaliphiles - Alkaline– Soda lakes in Africa and Western U.S.
• Halophiles - Highly saline– Natural salt lakes and manmade pools– Sometimes occurs with extreme
alkalinity
Survival
• Temperature extremes– Every part of microbe must
function at extreme• “Tough” enzymes for Thermophiles • “Efficient” enzymes for
Psychrophiles
–Many enzymes from these microbes are interesting
Life at High Temperatures, Thomas M. Brock
Survival
• Chemical extremes– Interior of cell is “normal”– Exterior protects the cell
• Acidophiles and Alkaliphiles sometimes excrete protective substances and enzymes
• Acidophiles often lack cell wall• Some moderate halophiles have high concs
of a solute inside to avoid “pickling”– Some enzymes from these microbes are
interesting
What are enzymes?• Definition - a protein that catalyses
(speeds up) chemical reactions without being changed
What are enzymes?
• Enzymes are specific– Lock and key analogy
Enzyme
Substrate A
Product B
Product C
What are enzymes?
• Activation energy– Enzymes allow reactions with lower
energyEn
erg
y
Time
Without Enzyme
With Enzyme
What are enzymes?• Enzymes are just a protein– They can be destroyed by
• Heat, acid, base– They can be inhibited by
• Cold, salt
• Heat an egg white or add vinegar to milk– Protein is a major component of
both- denatures
Practical Applications
• Extremozymes– Enzyme from Extremophile
– Industry & Medicine
• What if you want an enzyme to work – In a hot factory?– Tank of cold solution?– Acidic pond?– Sewage (ammonia)?–Highly saline solution?
One solution• Pay a genetic engineer to design a
“super” enzymes...– Heat resistant enzymes– Survive low temperatures– Able to resist acid, alkali and/or salt
• This could take years and lots of money
Extremophiles got there first
• Nature has already given us the solutions to these problems– Extremophiles have the enzymes
that work in extreme conditions
Endolithic algae from Antarctica; Hot springs in Yellowstone National Park, © 1998 Reston Communications, www.reston.com/astro/extreme.html
Thermophiles• Most interesting,
with practical applications
Many industrial processes involve high heat– 450C (113F) is a
problem for most enzymes
– First Extremophile found in 1972
Life at High Temperatures, Thomas M. Brock
PCR - Polymerase Chain Reaction
• Allows amplification of small sample of DNA using high temperature process– Technique is about 10 years old– DNA fingerprints - samples from crime
scene– Genetic Screening - swab from the mouth– Medical Diagnosis - a few virus particles
from blood • Thermus aquaticus or Taq
Life at High Temperatures, Thomas M. Brock
Psychrophiles• Efficient enzymes to work in the cold– Enzymes to work on foods that need to
be refrigerated– Perfumes - most don’t tolerate high
temperatures– Cold-wash detergents
Algal mats on an Antarctic lake bottom, © 1998 Reston Communications, www.reston.com/astro/extreme.html
Acidophiles
• Enzymes used to increase efficiency of animal feeds– enzymes help animals
extract nutrients from feed– more efficient and less
expensive
Life at High Temperatures, Thomas M. Brock
Alkaliphiles
• “Stonewashed” pants– Alkaliphilic enzymes soften fabric and
release some of the dyes, giving worn look & feel
• Detergents– Enzymes dissolve proteins or fats– Detergents do not inhibit alkaliphilic
enzymes
Halophiles• What is a halophile?• Diversity of Halophilic Organisms• Adptation Strategies
– Osmoregulation-“Compatible Solute” Strategy– “Salt-in” Strategy
• Interesting Facts and Applications
What is a halophile?
• Halophile = “salt loving; can grow in higher salt concentrations
• Based on optimal saline environments halophilic organisms can be grouped into three categories:– extreme halophiles, – moderate halophiles, and – slightly halophilic or halotolerant organisms
• Some extreme halophiles can live in solutions of 25 % salt; seawater = 2% salt
Diversity of Halophilic Organisms
• Halophiles are a broad group &t can be found in all three domains of life.
• Found in salt marshes, subterranean salt deposits, dry soils, salted meats, hypersaline seas, and salt evaporation ponds.
Unusual Habitats
• A Pseudomonas species lives on a desert plant in the Negev Desert- the plant leaves secretes salt through salt glands.
• A Bacillus species is found in the nasal cavities of desert iguanas- iguanas nasal cavities have salt glands which secrete KCl brine during osmotic stress.
Osmoregulation
• Halophiles maintain an internal osmotic potential that equals their external environment.
• Osmosis is the process in which water moves from an area of high concentration to an area of low concentration.
Osmoregulation
• In order for cells to maintain their water they must have an osmotic potential equal to their external environment.
• As salinity increases in the environment its osmotic potential decreases.
• If you placed a non halophilic microbe in a solution with a high amount of dissolved salts the cell’s water will move into the solution causing the cell to plasmolyze.
Osmoregulation
• Halophiles have adapted to life at high salinity in many different ways.– Structural modification of external
cell walls- posses negatively charged proteins on the outside which bind to positively charged sodium ions in their external environments & stabilizes the cell wall break down.
“Compatible Solute” Strategy
• Cells maintain low concentrations of salt in their cytoplasm by balancing osmotic potential with organic, compatible solutes.
• They do this by the synthesis or uptake of compatible solutes- glycerol, sugars and their derivatives, amino acids and their derivatives & quaternary amines such as glycine betaine.
• Energetically synthesizing solutes is an expensive process.– Autotrophs use between 30 to 90 molecules of
ATP to synthesize one molecule of compatible solute.
– Heterotrophs use between 23 to 79 ATP.
“Salt-in” Strategy• Cells can have internal concentrations
that are osmotically equivalent to their external environment.
• This “salt-in” strategy is primarily used by aerobic, extremely halophilic archaea and anaerobic bacteria.
• They maintain osmotically equivalent internal concentrations by accumulating high concentrations of potassium chloride.
“Salt-in” Strategy• Potassium ions enter the cell
passively via a uniporter. Sodium ions are pumped out. Chloride enters the cell against the membrane potential via cotransport with sodium ions.
• For every three molecules of potassium chloride accumulated, two ATP are hydrolyzed making this strategy more energy efficient than the “compatible solute” strategy.
“Salt-in” Strategy
• To use this strategy all enzymes and structural cell components must be adapted to high salt concentrations to ensure proper cell function.
Halobacterium: an extreme halophile
• Halobacterium are members of domain archaea.
• Widely researched for their extreme halophilism and unique structure.
• Require salt concentrations between 15% to saturation to live.
• Use the “salt-in” strategy.• Produce ATP by respiration or by
bacteriorhodopsin.
Halobacterium• May also have halorhodopsin
that pumps chloride into the cell instead of pumping protons out.
• The Red Sea was named after halobacterium that turns the water red during massive blooms.
Facts
• The term “red herring” comes from the foul smell of salted meats that were spoiled by halobacterium.
• There have been considerable problems with halophiles colonizing leather during the salt curing process.
Applications
• The extraction of carotene from carotene rich halobacteria and halophilic algae that can then be used as food additives or as food-coloring agents.
• The use of halophilic organisms in the fermentation of soy sauce and Thai fish sauce.
Applications
• Other possible applications being explored:– Increasing crude oil extraction
(MEOR)– Genetically engineering halophilic
enzymes encoding DNA into crops to allow for salt tolerance
– Treatment of waste water (petroleum)
Conclusions
• Halophiles are salt tolerant organisms.
• They are widespread and found in all three domains.
• The “salt-in” strategy uses less energy but requires intracellular adaptations. Only a few prokaryotes use it.
• All other halophiles use the “compatible solute” strategy that is energy expensive but does not require special adaptations.