tema 7: solute transportacademic.uprm.edu/~lrios/4368/tema7.pdf · tema 7: solute transport chapter...
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Tema 7: Solute Transport
Chapter 16Pages 417 - 437
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Water solublemolecules
Gram + bacteria
Permeabilitybarrier
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Water solublemolecules
Gram - bacteria
Permeabilitybarrier
Permeabilitybarrier
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This allows the bacterium to maintain an internal environment different from theexternal….
1- Metabolites could be maintained at an intracellular concentrationthat is orders of magnitude higher than the extracellularconcentration.
A- rapid enzymatic reactions.
B- retention of metabolic intermediates within the cell.
2- Minimizes the passive diffusion of ions.
A- maintain the electrochemical proton and sodiumion gradient.
1- ATP synthesis2- solute transport
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This implies that:
All water soluble molecules must enter and leave the cellthrough…
permease
transporter
carrier
porter
in
out
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How can we study transport?
Phospholipids+
water
sonicate buffer
Detergent-coated protein
liposomesproteoliposome
Cytochromeprotein
Lactose permease
Oxalate/formateantiporter Na+/H+
antiporter
Histidinepermease
Cytochromeoxidase
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proteoliposome
V max
V max
2
K m [S]
v
Vmax S
Km + Sv =
Michaelis-Menten constant
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liposome
proteoliposome
[S]
v
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Energy-dependent transport
Primary transport systems: driven by an energy-producing metabolic event,include proton translocation drive by ATP, light, or OX-RED Rxn.
1- Na ion transporting decarboxylases2- uptake of organic or inorganic solutes3- uptake of sugars
Secondary transport systems: driven by electrochemical gradients.
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ATP- binding cassette (ABC) transporters
2 subunits αβhydrophobic
2 subunits ααhydrophilic
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in outGram - cell
porinOMCM
ABC transporter
cytoplasm periplasm
ATP
ADP + Pi
sugars
amino acids
Small peptides
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ATP-drive K+ influx
Potassium is the principal cation in bacteria….. It play a role in:
Osmotic homeostasis, pH homeostasis, is a cofactor for many enzymes and ribosomes.
There are 2 transport systems:There are 2 transport systems:
1- TrK system (major route) is constitutively expressed and operatesat high rates but low affinity (Km ~ 2 mM).
2- Kdp system it depends on the salt-osmolarity of the medium, it has avery low Km ~ 2 µM
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The phosphotransferase system
Accumulates carbohydrates as the phosphorylated derivative.
This system is lacking in aerobes for the most part and it does not existin archaea and eukaryotes
This is a group translocation system!!! And not active transport.
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How to determine the source of energy for transport
Does it use ATP or ∆p??
We must do certain things before we ca determine this!!
1- inactivated the ATP synthase….How?
a- use ATP synthase mutants
b- DCCD (N,N’-diclohexylcorbodi-imide
2- perturbing the intracellular levels of ATP…..How?
a- starve the cell
b- add an inhibitor like arsenate (substrate level P)
c- to increase ATP, add glucose but limit respirationusing the inhibitor cyanide.
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3- perturbing the ∆p……How?
a- using Ionophores
B- addition of substrates that feed electrons to the e- transport chain
Drug-export systemThey are an important way by which bacteria become resistant to
antimicrobial agents. (antibiotics, dyes, detergents, disinfectants,and antiseptics.)
Two types: 1) dedicated and 2) multidrug
In M. tuberculosis, the TAP system pumps out both:1) aminoglycosides (gentamicin, kanamycin, neomycin, netilmicin, paromomycin, streptomycin, tobramycin and apramycin. 2) tetracycline
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Antimicrobial Agents and Chemotherapy, December 2000, p. 3249-3256, Vol. 44, No. 12
Lakshmi P. K. et.al.
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MDR system
Major classes: primary and secondary
The primary systems are energized by ATP hydrolysis.
They use an ABC transporter!
Membrane fusionProtein (AcrA)
The secondary systems are energized by proton antiport or sodium!
E. coli, N. gonorrhoea, N. meningitidis, V. cholerae,and V. parahaemolyticus.
Protein (AcrA)
porin
Efflux transpoter(AcrB)
OM channel(TolC)
AcrAB-TolCSystem pump