The Exotoxins of Escherichia coli

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The Exotoxins of Escherichia coli

• Professor Mark Pallen
• University of Birmingham

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The enterotoxins of enterotoxigenic E. coli

• plasmid-encoded
• Heat-labile toxins (LTs)
• Heat-stable toxins (STs)

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Heat -labile Toxins (LTs)

• Antigenic variants LT-I and LT-II
• LT-I neutralised by anti-cholera toxin
• LT-II not neutralised by anti-CT
• Importance in human disease uncertain

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LT-I

• AB5 toxin 88kDa
• A subunit 30 kDa
• Proteolytically knicked to give A1 and A2
• A1 enzymatically active
• A2 interacts with B subunits

• Five identical B subunits
• bind GM1 gangliosides
• Used as mucosal adjuvant
• LT holotoxin secreted to periplasmic space
• Mechanism of release across OMP uncertain

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Heat-Labile Enterotoxin (LT)

• Lunar lander modules
• single hexameric protein assembly as seen in the
  crystal structure of the LT AB5 holotoxin.
• Lunar surface
• the outer membrane of an intestinal epithelial
  cell.
• Protrusions from membrane under toxin
• 5 copies of saccharide component of ganglioside
  GM1 to which toxin binds.
• GM1 is a normal membrane component which the
  toxin co-opts as a receptor

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Heat-Labile Enterotoxin (LT)

• ADP-ribosylates membrane GTPase Gs
• regulates host cell adenylate cyclase, determines
  level of cAMP
• active (GTP-bound) Gs increases activity of
  adenylate cyclase
• GDP-bound form renders adenylate cyclase inactive
• active Gs normally produced following hormone
  stimulation, converted to inactive form after a
  short time
• ADP-ribosylation of Gs short-circuits off-on
  control by locking Gs in "on" form.

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LT-I
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LT-I

• LT-I Holotoxin binds cells, enters by endocytosis
• A1 subunit escapes vesicle
• A1 subunit is ADP-ribosyltransferase
• acting on Arg 201 of Gs protein on basolateral
  cell surface
• to cause permanent activation of adenylate
  cyclase
• raised cAMP levels cause water and electrolyte
  loss via CFTR

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LT-II

• Broadly similar to LT-I
• A subunits 57 amino-acid identity
• B subunits NOT related to LT-I
• Bind different gangliosides
• Role in disease uncertain

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Heat-stable toxins (STs)

• LMW cysteine-rich peptides with multiple
  disulphide bonds
• BUT two distinct groups (Sta, STb) according to
  sequence and mechanism

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Heat-stable toxinsSTa EAST-1

• STa
• 19-aa peptide derived from larger precursor
• Six cysteine residues
• Binds guanylate cyclase C leading to increased
  cGMP
• triggering water and electrolyte loss via CFTR
• EAST-1
• similar to STa
• in several pathotypes
• ETEC, EAEC
• Yersinia, Vibrio
• Carried on Tn4521

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STb

• 48-aa peptide, four cysteines
• formed from larger precursor
• uncertain mechanism and role

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Shiga-like toxin

• Enzymatic A chain in red
• Cell binding B chains (Pentamer) bind Gb3
• The A-subunits of Shiga toxin, the Shiga-like
  toxins (SLTs), and ricin inactivate eukaryotic
  ribosomes by enzymatically depurinating 28S rRNA

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Shiga-like toxin Genetics

• Phage encoded
• Induced on lysis
• iron-regulated, via fur
• Variants stx1 stx 2

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Shiga-like toxinAction

• Ribotoxic pro-inflammatory effect in gut
• Leads to haemorrhagic colitis
• Action at a distance
• transport through blood
• endothelium/kidney damage
• Haemolytic uraemic syndrome

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Haemolysin

• Important in virulence in UTI
• On PAI in UPEC
• Found in other pathotypes (EHEC)
• HlyA RTX toxin
• 11-17 C-terminal acidic Gly-rich repeats
• Bind Ca
• Forms beta super-helix
• Synthesised as pro-toxin, undergoes maturation
• HlyBATPase

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Haemolysin Action

• Cell subversion
• Triggers G-protein-dependent generation of
  inositol triphosphate
• Pro-inflammatory effects, apoptosis
• Pore formation
• Cell lysis and death
• 1.0nm asymmetric pores, probably monomeric
• 4 TMHs at N-terminus
• Triggered by Cainduced conformational changes
  at C-terminus

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E. coli haemolysin biogenesis
RfaH
hlyCABD
ops

• RfaH activation protein
• ops element
• Both also in F pilus, LPS, O-antigen, capsule
  loci
• Multi-component RNAP complex

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E. coli haemolysin biogenesis
FA-ACP
proHlyA
HlyB
HlyCD
TolC
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Cytolethal distending toxin