Life in the Extreme

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Life in the Extreme Professor Michael J. Danson
Pasteur (1822-1895)
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Edouard Chatton (1937) dichotomy
prokaryotes eukaryotes
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Carl Woese rRNA gene sequence comparisons to
create a phylogenetic tree
Bacteria Archaea
Archaea Thermophiles 55?C to 110?C Halophiles sat
urated NaCl Methanogens anaerobic and generate
CH4 Psychrophiles -2?C to 10?C Acidophiles,
alkaliphiles
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What are the molecular adaptations to life under
extreme conditions?
DNA
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Arthrobacter 15oC
Bacillus 25oC
Thermoplasma 55oC
Sulfolobus 80oC
Pyrococcus 100oC
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Citrate synthase inter-subunit helices
F
F
G
G
M
M
L
L
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Intersubunit ionic interactions in citrate
synthase
Bacillus Thermoplasma
Sulfolobus Pyrococcus (25oC)
(55oC) (80oC)
(100oC)
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• Active site
• - more accessible
• - polarised electrostatic surface

• Flexibility
• - high flexibility of small domain
• - two loops longer
• - fewer Pro in loops
• - more Pro in helices
• - more solvent-exposed hydrophobics

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Halophilic citrate synthases
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Global Industry Analysts, San José, CA,
USA Global enzyme sales in 2003 2,300
million
Detergents 790 million Food applications
630 million Agriculture-feed 380
million Textile processing 240 million
Predictions by 2010 160,000 million
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Thermostable Enzymes in Biotechnology
Thermostability Ease of recombinant
purification Longer shelf-life Temperature
solvent stable
Thermoactivity Active at high
temperatures Lower viscosity Improved
rate of mass transfer High S
P Lower risk of contamination
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• Polysaccharide degrading enzymes
• cellulases, amylases, pectinases, xylanases

• Detergent enzymes
• proteases, amylases, cellulases, lipases

• Oil industry

• Molecular biology
• DNA polymerases (Taq, Pfu, Vent)

• Enzymes for biocatalysis (eg pharmaceuticals)

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In 1998, Genencor commercialized an extremophile
enzyme Puradex HA
Puradax cellulase enzyme was derived from a new
Bacillus species found in the Rift Valley soda
lake of East Africa during an expedition in 1992.
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Antarctic phosphatase New England Biolabs
2005-06 catalogue
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Enzyme discovery
Environmental sample
Sample and culture
enrichment
Nucleic acid extraction

Metatranscriptomics Metagenomics mRNA
DNA
Gene enrichment
Genome enrichment
Gene discovery
Sequence-based screening Gene-specific PCR
Microarrays
Direct sequencing of libraries Sequence
assembly ORF assignment
Activity-based screening Expression screening
Primer walking or inverse PCR to obtain full
length gene
Gene product
Expression of ORFs in suitable host