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Microbiology and Molecular Biology for Engineers
IGEM, 20 June 2006
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There are three types of cell
• A: Archaea
• B: Bacteria (Gram positive, Gram negative)
• E: Eukarya (Animals, plants, yeasts, others)
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Eschericha coli• Kingdom/Division Proteobacteria• Class/Subdivision Gammaproteobacteria• Family Enterobacteriaceae
hull
sensors communications
propulsion
control systems power plant
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Cell envelope
• Typical of Gram negative bacteria
• Outer membrane: repels hydrophobic molecules.
• Peptidoglycan sacculus: resists osmotic pressure.
• Cell membrane: main permeability barrier.
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Energy generation• Energy generated by oxidation or disproportionation of
organic molecules.• Stored as non-equilibrium ATP/ADP ratio and trans-
membrane proton gradient.
Photon capture
Respiration
Fermentation
Trans-membrane proton gradient
ATP/ADP ratio
Motility
Transport processes
DNA, RNA andprotein synthesis
F1F0 ATPase
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Motility• Swimming motility: rotating flagella powered by proton influx.• Helical filaments: 20 nm diameter, 5 to 20 m long.
CM
PG
OML-ring
P-ring
MS-ring
hook
filament
motor proteinswitch protein
(Gram negative bacteria only)
EM of flagellar base structure
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Two-component sensor systems
• Sense external stimuli.• External stimulus causes modification of internal protein.
Cell membrane
CYTOPLASM
PERIPLASM
ligand
SensorKinase
PResponseRegulator
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Chemotaxis
• Swimming towards an attractant or away from a repellant is accomplished by a biased random walk – variable length runs interspersed with random changes of direction (tumbles).
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Mechanism of chemotaxis• Attractant / repellent chemicals are detected by
chemotaxis receptors (MCPs).• Phosphorylation state of CheY alters frequency of tumbles.• Methylation of MCPs decreases sensitivity.
MCPligand
CheW
CheA
CheY
CheY P
phosphatase
promotes tumbling
CheB
CheB Pincreases MCP sensitivity
phosphatase
Glu-Me
methylase
demethylase:
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Intercellular communication• Cells sense population density by
‘quorum sensing’.• Detect critical density of an
autoinducer, usually a homoserine lactone (LuxI/LuxR-type system)
luxC luxD luxA luxB luxEluxI luxGlux promoter
LuxR
OHHL
O
O
NO
O
Picture of squid
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Life cycle 1: shaken broth cultures
• Exponential growth phase followed by stationary phase with different genes expressed.
lag phase
exponentialphase
stationaryphase
declinephase
Biomass/Optical density
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Life Cycle 2: in nature
• Cells prefer to grow attached at solid-liquid interfaces (biofilm).
swimming cells
attached cells microcolonies mature biofilm
detachment
surface-associated motility
quorum sensing
swimming motility, chemotaxis
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Stochastic vs. mean field models
• Simplistic models often treat biomass as a single compartment.
• More realistically, billions of individual cells which may be in quite different states.
• Therefore, oscillators etc. must include a cell synchronization mechanism unless individual cells are to be monitored (eg by FACS or fluorescence microscopy).
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Hosts other than E. coli
• Salmonella: related to E. coli but better secretion of proteins into the medium. Problem: pathogen.
• Bacillus: good secretion of proteins, forms highly stable resting state (endospores).
• Budding yeast (Saccharomyces cerevisiae): eukaryotic cells.
• Plant, insect and mammalian cells.
endospores
yeast
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Coming up…
• What the stuff inside the cell is made of, and how it works.
• How to modify it.