Lecture 6' Microbial Adaptations 1

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Lecture 6. Microbial Adaptations 1

• Extremophilic micro-organisms
• Adaptive stresses
• Thermophiles
• Macromolecular stability
• Psychrophiles
• Halophiles
• Acidophiles

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Extremophiles
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Adaptive stresses

• Thermophiles
• Psychrophiles
• Halophiles
• Acidophiles
• Alkalophiles
• Barophiles
• Radiophiles
• Oligotrophy

• Denaturation and chemical destruction of macro-
  and small molecules membrane hyperfluidity
• Membrane hypofluidity Slow reaction kinetics
• Hyperosmotic stress
• Chemical stability membrane potentials
• Proton pumping against a negative pH gradient
• Unfavourable reaction equilibria reaction
  kinetics
• Molecular damage
• Nutrient deficiency and uptake affinity

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Thermophiles to Hyperthermophiles
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Thermophilic biotopes and organisms
Bacillus stearothermophilus Thermus
aquaticus Thermotoga spp. Sulfolobus spp.
Pyrococcus spp. Pyrodictium spp. Methanococcus
Terrestrial and deep sea hydrothermal systems
Methanococcus jannaschii
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Hyperthermophiles in the Tree of Life
BACTERIA
ARCHAEA
EUKARYA
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Living at high temperatures Problems and
Solutions

• Proteins denature at high temperatures.
• Many essential small molecules (e.g., ATP, NADH
  etc) are unstable at high temperatures.
• Membranes become more fluid at high temperatures.
• Reaction rates increase with temperature
  metabolic overrun?

• Thermodynamic stability of proteins is increased.
• Not known exactly metabolic channeling may be
  important.
• Membrane lipid compositions change to reduce
  fluidity.
• Enzyme conformational mobility and expression
  levels control metabolic rates

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Comparative stability of b-glycosidases
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Molecular mechanisms of protein stability

• Proteins are stabilised by weak intramolecular
  interactions (h? lt H-bonds lt ionic bonds) and
  protein-solvent interactions.
• Comparisons of 3D x-ray structures of homologous
  proteins from different thermal sources show
• gt Intramolecular packing
• lt External loops
• More helix-forming aas
• Stabilised helix dipoles
• gt Proline residues
• lt Asn residues
• Salt bridge networks

Higher protein stability
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Protein stability a balance between the net
free energy of the folded and denatured states

• N ? D model
• DG comprises enthalpy and entropy components
• Maximum enthalpy from intramolecular bonds
• Minimum entropy
• Folded proteins have high entropy
• Solvent entropy is minimised in folded state

DG Free energy of denaturation
Unfolded
Folded
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Stabilising lipids and reducing membrane fluidity

• Increase hydrophobic interactions between lipids
• Reduced fatty acid branching
• Increased fatty acid chain length
• Cyclisation
• Stabilise chemically sensitive ester bonds
• Ether-linked lipids in Archaea only
• Reduce lateral mobility of lipids
• C40 monolayer lipids in Archaea only

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Psychrophiles living at low temperatures
Arctic, Antarctic, deep Marine and alpine
regions
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Organisms at low temperatures

• Psychrophiles Topt, 15oC, Tmax lt 20oC, Tmin ,
  lt0oC
• Psychrotolerants (psychrotrophs) Topt, 20oC,
  Tmax gt 20oC, Tmin , gt3oC
• Very wide species diversity of Bacteria, Fungi,
  Algae
• E.g., Planococcus, Staphylococcus, Nostoc,
  Actinomyetes
• Limited diversity of Archaea
• Methanogens, uncultured crenarchaeota

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PsychrophilesProblems and Solutions

• Low metabolic rates
• Membrane rigidity
• Protein flexibility
• Cytoplasmic freezing

• Grow slowly
• Adapt membrane fluidity
• gt Unsaturation
• lt Chain length
• gt Methyl branching
• Increase conformational flexibility
• Reduce intramolecular bonding
• Accumulation of solutes (freeze prevention)
• Ice-nucleating proteins (freeze control)

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Extreme halophiles

• Living at very high salt concentrations
• Vertebrates lt 1.5M
• Halobacteria 1.5 3M
• Haloarchaea 3 5.2M

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Examples of moderate and extreme halophiles

• Bacteria
• Microccus
• Bacillus
• Flavobacterium
• Halomonas
• Vibrio
• Alteromonas
• Pseudomonas

• Eukaryotes
• Artemia
• Dunaliella
• Archaea
• Halobacterium
• Halorubrum
• Haloferax
• Haloarcula
• Halococcus

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Extreme halophiles in the tree of life
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Extreme halophilesProblems and Solutions

• Osmoregulation
• Protein stability

• Archaea
• Accumulation of intracellular salts (5M KCl)
• Bacteria
• Accumulation of low molecular weight solutes
  (osmolytes) with osmotic potential
• Increases in acidic a.a.s

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Osmolytes

• Glycerol (algae, fungi)
• Sugars, polyols and derivatives
• Glycine betaine
• a-amino acids (P, E)
• Ectoine, Hydroxyectoine
• Charged or hydroxylated compounds
• which H-bond with water molecules

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Protein stability in Halophiles

• High ionic strength (salt concentration)
  denatures proteins
• Dissociation of salt bridges
• Removal of protein water shell
• Halophilic proteins have increased surface
  charged amino acids
• Stabilised by K salt networks
• Increased water shell
• Many halophilic proteins require molar KCl for
  stability

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Acidophiles

• Geothermal areas
• Acid mine drainage

pH 0 3
2So 3 O2 2 H2O à 2 H2SO4
4 FeS2 15 O2 14 H2O à 4 Fe(OH)3 8 H2SO4
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Acidophiles are phylogenetically diverse

• Eukaryotes
• Fungi (numerous)
• Algae (Cyanidium)
• Protozoa (flagellates, ciliates, amoebae)
• Bacteria
• Heterotrophs (Alicyclobacillus, Acidiphilium,
  Sulfobacillus)
• Autotrophs (Acidithiobacillus)
• Archaea
• Numerous heterotrophs and autotrophs, many
  thermophilic (Thermoplasma, Sulfolobus,
  Acidianus, Metallosphaera)

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Chemoautotrophic metabolism in acidophiles

• Sulfolobus
• Acidianus
• Stygioglobus
• Aquifex
• Sulfobacillus

• Aerobic sulphur oxidation (So ? SO4-2 O2 ?
  H2O)
• Aerobic sulphide oxidation (S-2 ? SO4-2 O2 ?
  H2O)
• Anaerobic hydrogen oxidation (H2 So ? H2S)
• Aerobic hydrogen oxidation (H2 1/2O2 ? H2O)
• Aerobic iron oxidation (FeII ? FeIII O2
  ? H2O)

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AcidophilesProblems and Solutions

• pH homeostasis
• Molecular stability

• Intracellular pH trans-membrane potentials
• Internal pH is maintained at 5-7 by maintained
  by proton pumping high external proton drives
  chemiosmotic ATP synthesis
• Acid stable proteins
• Stabilised by increase in charged amino acids