INF1 - F. TOXIN TARGETS AND SEPSIS-COVERED

Question 1

how do bacteria make us feel ill after infection

Answer 1

• release toxins that weaken immune system and cause diarrhoea and autonomic dysfunction (heart rate or blood pressure)
• immunological response to toxins can lead to symptoms of illness: fever and shock

Question 2

what are exotoxins

Answer 2

• secreted proteins (enzymes and accessory proteins) secreted from living bacteria that attack host cells
• specific molecular targets
• high toxicity

Question 3

examples of exotoxins

Answer 3

ADP-ribosyltransferases
Monoglycosyltransferases
Metalloproteases

Question 4

what bacteria mainly releases exotoxins

Answer 4

anaerobic gram positive bacteria - especially Clostridium species
ie - botulism, whooping cough, cholera, dysentry

Question 5

what are the 3 classes of exotoxin

Answer 5

1. enzyme mechanism (ie, by the type of enzyme they secrete)
   - ADP-ribosyltransferases, monoglycosyltransferases
   metalloproteases
2. cellular target
   - neurotoxins, cytotoxins, enterotoxins
3. molecular target
   - synaptic release apparatus
   - GPCRs
   - cytoskeleton
   - protein synthesis apparatus

Question 6

what are the 2 main neurotoxins

Answer 6

1. botulinum toxin produced by clostridium botulinum
   - Zinc metalloproteases that cleave SNARE proteins
   - degrades vesicle release machinery
   - prevents fusion of vesicle with membrane
   - BoNT acts on ACh releasing cells
2. tetanus toxin produced by clostridium tetani
   - same enzymatic mechanism as BoNT
   - TeNT acts on glycine and GABA releasing cells

Question 7

what is a SNARE complex

Answer 7

mediates fusion of synaptic vesicles with the presynaptic plasma membrane resulting in exocytosis of neurotransmitters

Question 8

what 2 toxins target GPCRs

Answer 8

1. cholera toxin produced by vibrio cholerae
   - ADP-ribosyltransferase adds ADP-ribose moiety to alpha-s subunits of heterotrimeric G proteins
   - persistent activation of adenylyl cyclase by activation of Gs
2. pertussis toxin produced by bordetella pertussis (whooping cough)
   - ADP-ribosylation of alpha-i subunits of heterotrimeric G proteins
   - persistent activation of adenylyl cyclase by activation of Gi

Question 9

pharmacology of cholera toxin

Answer 9

normally: GDP substituted with GTP and G alpha-s binds to AC and releases cAMP

when toxin binds: targets G alpha-s, causes ribosylation and causes G alpha-s to be permanently switched on and cAMP continually produced

Question 10

pharmacology of pertussis toxin

Answer 10

normally: GDP substituted with GTP, G alpha-i binds to AC, AC is switched off and prevents cAMP being produced

when toxin binds: binds to inactive form Gi-coupled receptor and keeps receptor in inactive state, switches off ‘off’ switch so cAMP continually produced

Question 11

what toxins target cytoskeleton in the cell

Answer 11

numerous clostridium sp. toxins
- ADP-ribosylation of monomeric G-actin
- inhibits actin polymerisation
- disrupts cell morphology, division, transport of organelles etc

clostridium difficile toxins A and B
- glycosyltransferases: use UDP-glucose to transfer glucose moiety to Rho family of small GTPases
- inhibits actin polymerisation (+ many other signalling cascades)

Question 12

pharmacology of disruption of actin polymerisation

Answer 12

F-actin is a polymer of individual G-actin monomers and polymerisation is promoted by Rho proteins (Rho-GTPase)

C. perfringens toxin causes ADP ribsosylation of G-actin
This prevents G-actin from forming F-actin polymer
Depletes pool of monomers

C. difficile (processed) catalyses transfer of glucose from UDP-glucose and it binds to Rho protein and inactivates it. Can’t have polymerisation without Rho-GTPase so can’t get F-actin formation

Question 13

what 2 toxins target protein synthesis

Answer 13

1. diphtheria toxin produced by corynebacterium diphtheriae
   - ADP-ribosylation of elongation factor 2
   - inhibition of translation
2. shiga toxin produced by shigella dysenteriae
   - N-glycosidase: hydrolyses the 28S RNA in the ribosome

Question 14

what are endotoxins

Answer 14

• structural macromolecules (lipids, polysaccharides) released where bacteria are destroyed (ie - from cell wall)
• gram -ve
• non-specific
• low toxicity but immunogenic
• not released intentionally

Question 15

what is the main endotoxin

Answer 15

LPS
Lipid A contributes toxicity
Polysaccharides (O and R) contribute immunogenicity

LPS activates signalling through Toll-like receptors (TLR4) in immune cells (macrophages, monocytes) - polysaccharide recognised as an epitope by immune system

This triggers release of cytokines and activation of complement cascade

Question 16

what is a cytokine storm

Answer 16

LPS-induced immune response can be excessive
Cytokines, nitric oxide, the pro-infam mediators produced
If response spreads beyond site of infection, systemic inflam response - sepsis

Question 17

how does sepsis occur

Answer 17

• pro-inflam cytokines and procoaggulants release by macrophage and
  vasodilators produced by macrophage increase lumen of smooth vessels and hence a big decrease in blood pressure
• the endothelial wall of blood vessels starts to form gaps in between each cell which causes leakage - haemorrhage
  organ failure as not enough blood supply