Lecture 9 BIOL 533 1
Bacterial Toxins
BIOL 533
Lecture 9
Medical Microbiology
Lecture 9 BIOL 533 2
Bacterial Toxins,
General Aspects
? Definition
– Soluble substances that alter normal
metabolism of host cells with deleterious
effects on the host
? Host range
– Known for bacteria,but possible that they
play a role in diseases caused by fungi,
protozoa,and worms
Lecture 9 BIOL 533 3
Bacterial Toxins,
General Aspects
? Toxin type
– Exotoxin—protein produced by bacteria either
excreted or bound to bacterial surface and
released when lysed
– Endotoxin—lps of the outer membrane of
Gram— bacteria
? Acts as toxin only under special circumstances
Lecture 9 BIOL 533 4
Bacterial Toxins,
General Aspects
? Specificity
– Some act on certain cell types
– Other affect wide range of cells and tissues
? Numbers produced by single bacterium
– Some produce none
? Pneumococci
Lecture 9 BIOL 533 5
Is Toxin Important in Infection?
? Questions to ask,
– Is virulence quantitatively correlated with
toxin production?
– Does the purified toxin produce damage?
– Can a specific antibody (antitoxin) prevent or
alleviate the manifestations of the disease?
– If toxin production is impaired by a mutation,
is the disease process affected?
Lecture 9 BIOL 533 6
If So,What are Toxin Properties?
? Questions to ask,
– What is the mechanism of action?
– Why is it specific for certain cells or tissues?
– Does the pathogen make other toxins,and if
so,do they interact with one another?
? Some make none,pneumococci
? Some make only one,agents that cause cholera,
diphtheria,tetanus,and botulism
? Other make many,staphylococci,streptococci
Lecture 9 BIOL 533 7
Toxin Production
? Properties
– Dispensable,but essential under certain
situations where survival and spread are at
stake
– Genes frequently carried on plasmids and
temperate bacteriophage
Lecture 9 BIOL 533 8
Toxin Production
– Found on phage; toxin genes for,
? Diphtheria
? Botulism
? Scarlet fever
? Toxic streptococci (―flesh-eating‖)
– Found on plasmids,
? E,coli toxin causes diarrhea
? S,aureus toxin causes ―scalded skin syndrome‖
? E,coli 0157:H7
Lecture 9 BIOL 533 9
Toxin Production
– Properties
? Mobile elements ensure that genes can be spread
to nontoxigenic derivatives or be lost from cell
– Experimentally called ―curing‖—get nontoxigenic
derivatives
– Phase of production
? Some produced continuously by growing bacteria
? Other synthesized when cells enter stationary
phase (true also for many antibiotics)
Lecture 9 BIOL 533 10
Toxin Production
– Explanation
? Certain toxins may help bacteria get scarce
nutrients
? Example,high levels of diphtheria toxin produced
when cell depleted of iron
– Very little free iron in normal tissue
– Is this a way for organisms to obtain it from dead tissue?
Lecture 9 BIOL 533 11
Toxin Production
– Sporulating bacteria sometimes release toxins
during spore formation
? Bacterial cells eventually lyse and liberate
cytoplasmic proteins,including toxins
– Examples,organisms that cause botulism,gas gangrene,
or tetanus
? In contaminated wound,some organisms are
growing and some are sporulating
– End result is continual production
Lecture 9 BIOL 533 12
Mechanism of Action
? General aspects
– Sphere of influence
? Some act locally,killing wbc nearby
? Others help organism to spread in host itssues by
degrading connective tissue
? Still others are disseminated very far from site
where synthesized
– Diphtheria toxin made in throat,but acts on heart and
brain
Lecture 9 BIOL 533 13
Mechanism of Action
? Level of toxicity
– Work at extremely low levels; include
strongest poisons known
? 1 g tetanus,botulinus,or Shiga toxin is enough to
kill 10 million people
? 100-fold more is required for diptheria
? 1000-fold more for Pseudomonas A
Lecture 9 BIOL 533 14
Mechanism of Action
? Mechanisms of damage
– Lysis of host cells
– Stop or interfere with cell growth
– Exaggerate normal physiological mechanisms
? By depressing or augmenting particular functions,
toxins can kill without damaging any cells
– Tetanus toxin paralyzes body without affecting target
neurons
– Cholera toxin speeds up normal excretory process,
resulting in massive loss of water
Lecture 9 BIOL 533 15
Mechanism of Action
? Toxins that assist bacterial spread in
tissues
– Properties
? Do not target any type of cell
? Include degradative enzymes that allow spreading
Lecture 9 BIOL 533 16
Mechanism of Action
? Examples,
– Streptococci
? Some secrete
– Hyaluronidase—breaks down hyaluronic acid (connective
tissue)
– DNase—thins out pus made viscous by DNA from dead
white blood cell
– Streptokinase (protease)—cleaves precursor of
plasminogen activator to active form
? Converts plasminogen to plasmin (serum protease
that dissolves fibrin clots)
Lecture 9 BIOL 533 17
Mechanism of Action
? Examples,
– Similar roles suggested for elastases and
collagenases of other organisms
? In this case,are unregulated forms of enzymes
that also exist in uninfected host (activity is
normally under control)
Lecture 9 BIOL 533 18
Mechanism of Action
? Toxins that lyse cells
– General aspects
? Large class kill host cells by destroying their
membranes; act as lipases
? Example of lipase type,Clostridium perfringens
(gas gangrene) lecithinase
– Lyses cells indiscriminately because phosphatidylcholine
(lecithin) is ubiquitous in mammalian membranes
– Also hemolysins are of this type; lyse both red blood cells
and white blood cells
Lecture 9 BIOL 533 19
Mechanism of Action
? Act by inserting themselves in membrane forming
pores
– Mechanism,make membrane more permeable,water
pours into cytoplasm,cell begins to swell,and eventually
bursts
– At very low concentrations (not enough to cause lysis),
cell functions may be severely damaged,Slight
perturbations of permeability cause,
? Leakage of potassium ions needed for protein
synthesis and cell viability
? Low levels inhibit phagocyte functioning
Lecture 9 BIOL 533 20
Mechanism of Action
– Examples,
? Staphylococci ?-toxin (homogeneous pore former)
– Receptors exist—cells show 100-fold range in sensitivity
– Consequences of action,aggregation of platelets and
narrowing of blood vessels leads to necrosis
Lecture 9 BIOL 533 21
Mechanism of Action
– Examples,
? Streptococcal streptolysin 0 (heterogeneous pore
former)
– Binds to cholesterol in cell membrane
? Free toxin can be inactivated by cholesterol,but
once bound by membrane,it is impervious
– Consequences of the action,lyses red blood cells,but
not neutrophils or macrophage
? White blood cells are killed by low levels of toxin
because it acts preferentially on membranes of
lysosomes,releasing hydrolytic enzymes
Lecture 9 BIOL 533 22
Mechanism of Action
? Toxins that block protein synthesis
– Structure and mode of action
? Toxins that work outside the cell are variable in
structure and mode of action
? Toxins that work inside have a number of
similarities
Lecture 9 BIOL 533 23
Mechanism of Action
? Similarities
– Most have two portions (A-B toxins)
? Subunits
– Toxic activity (A)
– Binding to cell membrane (B)
– Can be one polypeptide chain or many
– Binding to membrane may be followed by
receptor-mediated endocytosis and
internaliztion of the toxin (some investigators
propose direct passage through pore)
Lecture 9 BIOL 533 24
Mechanism of Action
– ―A‖ moity is often latent,even after
engulfment
? May be activated by proteoytic cleavage and
reduction of disulfide bridges
– Toxins of diphtheria,cholera,tetanus,and Shigella are
synthesized as inactive precusors
Lecture 9 BIOL 533 25
Mechanism of Action
– May have common mode of action
? Catalyze transfer of adenosine-diphosphate group
from NAD to target proteins
? Examples of ADP-ribosyltransferases—toxins of,
– Diphtheria
– Cholera
– Exotoxin A (Pseudomonas aeruginosa)
Lecture 9 BIOL 533 26
Diphtheria Toxin
? How does toxin enter cell?
– A and B are single polypeptide chain
? Hydrophobic B portion binds to receptor on
membrane
? By this time,molecule is cleaved at sensitive site
between A and B portions,but is still covalently
associated by disulfide linkage
? Entire receptor-toxin complex enters cell by
receptor-mediated endocytosis
Lecture 9 BIOL 533 27
Diphtheria Toxin
– Once toxin enters,reduction S-S bond
separates A and B portion
– Acidic conditions within endosomal vesicles
promote insertion of B chain into endosomal
membrane
? Somehow,this facilitates passage of A into cytosol
– Resistant to denaturation and is long-lived within cells
– Accounts in part for potency (single molecule can kill
cell)
Lecture 9 BIOL 533 28
Diphtheria Toxin
? Mechanism of killing
– ADP-ribosylation of EF2 (protein that catalyzes
hydrolysis of GTP that drives movement of
ribosomes on eucaryotic mRNA)
? Reaction is,
EF-2 + NAD+? ADPR-EF2 + H+
Lecture 9 BIOL 533 29
Diphtheria Toxin
? EF2 is only known substrate for diphtheria toxin
– EF2 contains rare modification of one of histidine
residues and this is site recognized by toxin
– Mutant cells that cannot modify site are resistant
? Addition of ADP-ribose inactivates EF2
– Kills cells by irreversible block of protein synthesis
– P,aeruginosa exotoxin A works same as
diphtheria toxin
Lecture 9 BIOL 533 30
Mechanism of Action
? Phamacological toxins (elevation of cAMP-
cholera)
– Excess of cAMP interferes with phagocyte
functioning (chemotaxis and phagocytosis)
– Methods of increasing,
? Secretion of cAMP
? Secretion of adenyl cyclase to make more cAMP
? Secretion of toxin alters activity of host adenyl
cyclase (cholera)
Lecture 9 BIOL 533 31
Cholera Toxin
? Target tissue is small intestine epithelium
? Structure and mechanism of toxin
– Toxin has separate A and B subunits
? B has affinity for intestinal epithelial mucosa
? A ADP-ribosylates GTPase (part of complex that
makes cAMP)
– Synthesis of cAMP becomes unregulated; made in large
amounts
– Provokes loss of fluids and copious diarrhea
Lecture 9 BIOL 533 32
Cholera Toxin
– Structure of subunits
? Five B subunits and one A subunit
? A subunit is synthesized as single chain
– Then,after secretion,cleaved into two fragments (A1
and A2; held together by disulfide bonds)
Lecture 9 BIOL 533 33
Cholera Toxin
– Mechanism
? Whole toxin binds to 5 ganglioside receptors on
surface of intestinal epithelial cells
? A1-A2 portion enters cell and is cleaved into A1
and A2 pieces (by reduction of disulfide bonds)
– A1 fragment in enzymatically active
Lecture 9 BIOL 533 34
Cholera Toxin
– Regulation
? Normal
– Adenylate cyclase complex is membrane bound and is
composed of three proteins (Gs,R,cyclase)
– Gs protein is GTPase protein with two conformational
states
? Binds GTP—stimulates adenyl cyclase to make cAMP
? GTPase that cleaves GTP to GDP
Lecture 9 BIOL 533 35
Cholera Toxin
? Balance is determined by binding of R protein
– Binding of GTP by Gs stimulated by binding R protein
– R is receptor for several different hormones (adenergics)
– Whole picture—when R protein binds with hormone,
interacts with Gs protein to increase its binding of GTP
? Gs remains in active state to stimulate adenyl cyclase
Lecture 9 BIOL 533 36
Cholera Toxin
? Abnormal (cholera) normal action of R protein
mimicked by cholera toxin
– Promotes active state of Gs protein by different
mechanism
– ADP-ribosylates Gs at one of its arginine residues (Gs
protein locked into active conformation)
Lecture 9 BIOL 533 37
Mechanism of Action
? Other toxins that activate adenylate
cyclase
– Number of enterotoxins that produce diarrhea
? LT (labile)—E,coli
– Bordetella pertussis adenylate cyclase
? Raise level cAMP in leucocytes
Lecture 9 BIOL 533 38
Mechanism of Action
? Toxins that block nerve function
– Most lethal toxins known are tetanus and
botulinum toxins
? Tetanus toxin produces irreversible muscle
contraction
? Botulinum toxin blocks muscle contraction
Lecture 9 BIOL 533 39
Mechanism of Action
? General mechanism of both
– Consist of single polypeptide chains with A
and B regions
– Binding to ganglioside receptors specific for
nerve tissue
– Activated by proteolysis and disulfide
reduction,and they function intracellularly
Lecture 9 BIOL 533 40
Tetanus Toxin
? Acts at distance from central nervous
system
– Once bound to cell membranes,toxin is
internalized probably by receptor-mediated
endocytosis
Lecture 9 BIOL 533 41
Tetanus Toxin
– Transported through axonal processes to the
spinal cord
? Toxin interferes with synaptic transmission by
preferentially inhibiting release of
neurotransmitter,such as glycine from inhibitory
interneurons
? Excitory and inhibitory effects of motor neurons
become increasingly unbalanced,causing rigid
muscle contractions
– Cause of inhibitory synapse action unknown
Lecture 9 BIOL 533 42
Botulinum Toxin
? General aspects
– Intoxication,not infection; organism not
needed after toxin produced
– Toxin not destroyed by proteases of digestive
tract; probably complexed with other proteins
? Mechanism
– Affects peripheral nerve endings
Lecture 9 BIOL 533 43
Botulinum Toxin
– Once across the gut,it is carried in the blood
to neuromuscular junctions
? Bind to gangliosides at motor nerve endpoints and
is taken up by cell
? Subsequent events unknown
– Result in presynaptic block of release of acetylcholine
– Interruptions in nerve stimulation causes irreversible
relaxation of muscles—leads to respiratory arrest
Lecture 9 BIOL 533 44
Lecture 9
? Questions?
? Comments?
? Assignments..,
Bacterial Toxins
BIOL 533
Lecture 9
Medical Microbiology
Lecture 9 BIOL 533 2
Bacterial Toxins,
General Aspects
? Definition
– Soluble substances that alter normal
metabolism of host cells with deleterious
effects on the host
? Host range
– Known for bacteria,but possible that they
play a role in diseases caused by fungi,
protozoa,and worms
Lecture 9 BIOL 533 3
Bacterial Toxins,
General Aspects
? Toxin type
– Exotoxin—protein produced by bacteria either
excreted or bound to bacterial surface and
released when lysed
– Endotoxin—lps of the outer membrane of
Gram— bacteria
? Acts as toxin only under special circumstances
Lecture 9 BIOL 533 4
Bacterial Toxins,
General Aspects
? Specificity
– Some act on certain cell types
– Other affect wide range of cells and tissues
? Numbers produced by single bacterium
– Some produce none
? Pneumococci
Lecture 9 BIOL 533 5
Is Toxin Important in Infection?
? Questions to ask,
– Is virulence quantitatively correlated with
toxin production?
– Does the purified toxin produce damage?
– Can a specific antibody (antitoxin) prevent or
alleviate the manifestations of the disease?
– If toxin production is impaired by a mutation,
is the disease process affected?
Lecture 9 BIOL 533 6
If So,What are Toxin Properties?
? Questions to ask,
– What is the mechanism of action?
– Why is it specific for certain cells or tissues?
– Does the pathogen make other toxins,and if
so,do they interact with one another?
? Some make none,pneumococci
? Some make only one,agents that cause cholera,
diphtheria,tetanus,and botulism
? Other make many,staphylococci,streptococci
Lecture 9 BIOL 533 7
Toxin Production
? Properties
– Dispensable,but essential under certain
situations where survival and spread are at
stake
– Genes frequently carried on plasmids and
temperate bacteriophage
Lecture 9 BIOL 533 8
Toxin Production
– Found on phage; toxin genes for,
? Diphtheria
? Botulism
? Scarlet fever
? Toxic streptococci (―flesh-eating‖)
– Found on plasmids,
? E,coli toxin causes diarrhea
? S,aureus toxin causes ―scalded skin syndrome‖
? E,coli 0157:H7
Lecture 9 BIOL 533 9
Toxin Production
– Properties
? Mobile elements ensure that genes can be spread
to nontoxigenic derivatives or be lost from cell
– Experimentally called ―curing‖—get nontoxigenic
derivatives
– Phase of production
? Some produced continuously by growing bacteria
? Other synthesized when cells enter stationary
phase (true also for many antibiotics)
Lecture 9 BIOL 533 10
Toxin Production
– Explanation
? Certain toxins may help bacteria get scarce
nutrients
? Example,high levels of diphtheria toxin produced
when cell depleted of iron
– Very little free iron in normal tissue
– Is this a way for organisms to obtain it from dead tissue?
Lecture 9 BIOL 533 11
Toxin Production
– Sporulating bacteria sometimes release toxins
during spore formation
? Bacterial cells eventually lyse and liberate
cytoplasmic proteins,including toxins
– Examples,organisms that cause botulism,gas gangrene,
or tetanus
? In contaminated wound,some organisms are
growing and some are sporulating
– End result is continual production
Lecture 9 BIOL 533 12
Mechanism of Action
? General aspects
– Sphere of influence
? Some act locally,killing wbc nearby
? Others help organism to spread in host itssues by
degrading connective tissue
? Still others are disseminated very far from site
where synthesized
– Diphtheria toxin made in throat,but acts on heart and
brain
Lecture 9 BIOL 533 13
Mechanism of Action
? Level of toxicity
– Work at extremely low levels; include
strongest poisons known
? 1 g tetanus,botulinus,or Shiga toxin is enough to
kill 10 million people
? 100-fold more is required for diptheria
? 1000-fold more for Pseudomonas A
Lecture 9 BIOL 533 14
Mechanism of Action
? Mechanisms of damage
– Lysis of host cells
– Stop or interfere with cell growth
– Exaggerate normal physiological mechanisms
? By depressing or augmenting particular functions,
toxins can kill without damaging any cells
– Tetanus toxin paralyzes body without affecting target
neurons
– Cholera toxin speeds up normal excretory process,
resulting in massive loss of water
Lecture 9 BIOL 533 15
Mechanism of Action
? Toxins that assist bacterial spread in
tissues
– Properties
? Do not target any type of cell
? Include degradative enzymes that allow spreading
Lecture 9 BIOL 533 16
Mechanism of Action
? Examples,
– Streptococci
? Some secrete
– Hyaluronidase—breaks down hyaluronic acid (connective
tissue)
– DNase—thins out pus made viscous by DNA from dead
white blood cell
– Streptokinase (protease)—cleaves precursor of
plasminogen activator to active form
? Converts plasminogen to plasmin (serum protease
that dissolves fibrin clots)
Lecture 9 BIOL 533 17
Mechanism of Action
? Examples,
– Similar roles suggested for elastases and
collagenases of other organisms
? In this case,are unregulated forms of enzymes
that also exist in uninfected host (activity is
normally under control)
Lecture 9 BIOL 533 18
Mechanism of Action
? Toxins that lyse cells
– General aspects
? Large class kill host cells by destroying their
membranes; act as lipases
? Example of lipase type,Clostridium perfringens
(gas gangrene) lecithinase
– Lyses cells indiscriminately because phosphatidylcholine
(lecithin) is ubiquitous in mammalian membranes
– Also hemolysins are of this type; lyse both red blood cells
and white blood cells
Lecture 9 BIOL 533 19
Mechanism of Action
? Act by inserting themselves in membrane forming
pores
– Mechanism,make membrane more permeable,water
pours into cytoplasm,cell begins to swell,and eventually
bursts
– At very low concentrations (not enough to cause lysis),
cell functions may be severely damaged,Slight
perturbations of permeability cause,
? Leakage of potassium ions needed for protein
synthesis and cell viability
? Low levels inhibit phagocyte functioning
Lecture 9 BIOL 533 20
Mechanism of Action
– Examples,
? Staphylococci ?-toxin (homogeneous pore former)
– Receptors exist—cells show 100-fold range in sensitivity
– Consequences of action,aggregation of platelets and
narrowing of blood vessels leads to necrosis
Lecture 9 BIOL 533 21
Mechanism of Action
– Examples,
? Streptococcal streptolysin 0 (heterogeneous pore
former)
– Binds to cholesterol in cell membrane
? Free toxin can be inactivated by cholesterol,but
once bound by membrane,it is impervious
– Consequences of the action,lyses red blood cells,but
not neutrophils or macrophage
? White blood cells are killed by low levels of toxin
because it acts preferentially on membranes of
lysosomes,releasing hydrolytic enzymes
Lecture 9 BIOL 533 22
Mechanism of Action
? Toxins that block protein synthesis
– Structure and mode of action
? Toxins that work outside the cell are variable in
structure and mode of action
? Toxins that work inside have a number of
similarities
Lecture 9 BIOL 533 23
Mechanism of Action
? Similarities
– Most have two portions (A-B toxins)
? Subunits
– Toxic activity (A)
– Binding to cell membrane (B)
– Can be one polypeptide chain or many
– Binding to membrane may be followed by
receptor-mediated endocytosis and
internaliztion of the toxin (some investigators
propose direct passage through pore)
Lecture 9 BIOL 533 24
Mechanism of Action
– ―A‖ moity is often latent,even after
engulfment
? May be activated by proteoytic cleavage and
reduction of disulfide bridges
– Toxins of diphtheria,cholera,tetanus,and Shigella are
synthesized as inactive precusors
Lecture 9 BIOL 533 25
Mechanism of Action
– May have common mode of action
? Catalyze transfer of adenosine-diphosphate group
from NAD to target proteins
? Examples of ADP-ribosyltransferases—toxins of,
– Diphtheria
– Cholera
– Exotoxin A (Pseudomonas aeruginosa)
Lecture 9 BIOL 533 26
Diphtheria Toxin
? How does toxin enter cell?
– A and B are single polypeptide chain
? Hydrophobic B portion binds to receptor on
membrane
? By this time,molecule is cleaved at sensitive site
between A and B portions,but is still covalently
associated by disulfide linkage
? Entire receptor-toxin complex enters cell by
receptor-mediated endocytosis
Lecture 9 BIOL 533 27
Diphtheria Toxin
– Once toxin enters,reduction S-S bond
separates A and B portion
– Acidic conditions within endosomal vesicles
promote insertion of B chain into endosomal
membrane
? Somehow,this facilitates passage of A into cytosol
– Resistant to denaturation and is long-lived within cells
– Accounts in part for potency (single molecule can kill
cell)
Lecture 9 BIOL 533 28
Diphtheria Toxin
? Mechanism of killing
– ADP-ribosylation of EF2 (protein that catalyzes
hydrolysis of GTP that drives movement of
ribosomes on eucaryotic mRNA)
? Reaction is,
EF-2 + NAD+? ADPR-EF2 + H+
Lecture 9 BIOL 533 29
Diphtheria Toxin
? EF2 is only known substrate for diphtheria toxin
– EF2 contains rare modification of one of histidine
residues and this is site recognized by toxin
– Mutant cells that cannot modify site are resistant
? Addition of ADP-ribose inactivates EF2
– Kills cells by irreversible block of protein synthesis
– P,aeruginosa exotoxin A works same as
diphtheria toxin
Lecture 9 BIOL 533 30
Mechanism of Action
? Phamacological toxins (elevation of cAMP-
cholera)
– Excess of cAMP interferes with phagocyte
functioning (chemotaxis and phagocytosis)
– Methods of increasing,
? Secretion of cAMP
? Secretion of adenyl cyclase to make more cAMP
? Secretion of toxin alters activity of host adenyl
cyclase (cholera)
Lecture 9 BIOL 533 31
Cholera Toxin
? Target tissue is small intestine epithelium
? Structure and mechanism of toxin
– Toxin has separate A and B subunits
? B has affinity for intestinal epithelial mucosa
? A ADP-ribosylates GTPase (part of complex that
makes cAMP)
– Synthesis of cAMP becomes unregulated; made in large
amounts
– Provokes loss of fluids and copious diarrhea
Lecture 9 BIOL 533 32
Cholera Toxin
– Structure of subunits
? Five B subunits and one A subunit
? A subunit is synthesized as single chain
– Then,after secretion,cleaved into two fragments (A1
and A2; held together by disulfide bonds)
Lecture 9 BIOL 533 33
Cholera Toxin
– Mechanism
? Whole toxin binds to 5 ganglioside receptors on
surface of intestinal epithelial cells
? A1-A2 portion enters cell and is cleaved into A1
and A2 pieces (by reduction of disulfide bonds)
– A1 fragment in enzymatically active
Lecture 9 BIOL 533 34
Cholera Toxin
– Regulation
? Normal
– Adenylate cyclase complex is membrane bound and is
composed of three proteins (Gs,R,cyclase)
– Gs protein is GTPase protein with two conformational
states
? Binds GTP—stimulates adenyl cyclase to make cAMP
? GTPase that cleaves GTP to GDP
Lecture 9 BIOL 533 35
Cholera Toxin
? Balance is determined by binding of R protein
– Binding of GTP by Gs stimulated by binding R protein
– R is receptor for several different hormones (adenergics)
– Whole picture—when R protein binds with hormone,
interacts with Gs protein to increase its binding of GTP
? Gs remains in active state to stimulate adenyl cyclase
Lecture 9 BIOL 533 36
Cholera Toxin
? Abnormal (cholera) normal action of R protein
mimicked by cholera toxin
– Promotes active state of Gs protein by different
mechanism
– ADP-ribosylates Gs at one of its arginine residues (Gs
protein locked into active conformation)
Lecture 9 BIOL 533 37
Mechanism of Action
? Other toxins that activate adenylate
cyclase
– Number of enterotoxins that produce diarrhea
? LT (labile)—E,coli
– Bordetella pertussis adenylate cyclase
? Raise level cAMP in leucocytes
Lecture 9 BIOL 533 38
Mechanism of Action
? Toxins that block nerve function
– Most lethal toxins known are tetanus and
botulinum toxins
? Tetanus toxin produces irreversible muscle
contraction
? Botulinum toxin blocks muscle contraction
Lecture 9 BIOL 533 39
Mechanism of Action
? General mechanism of both
– Consist of single polypeptide chains with A
and B regions
– Binding to ganglioside receptors specific for
nerve tissue
– Activated by proteolysis and disulfide
reduction,and they function intracellularly
Lecture 9 BIOL 533 40
Tetanus Toxin
? Acts at distance from central nervous
system
– Once bound to cell membranes,toxin is
internalized probably by receptor-mediated
endocytosis
Lecture 9 BIOL 533 41
Tetanus Toxin
– Transported through axonal processes to the
spinal cord
? Toxin interferes with synaptic transmission by
preferentially inhibiting release of
neurotransmitter,such as glycine from inhibitory
interneurons
? Excitory and inhibitory effects of motor neurons
become increasingly unbalanced,causing rigid
muscle contractions
– Cause of inhibitory synapse action unknown
Lecture 9 BIOL 533 42
Botulinum Toxin
? General aspects
– Intoxication,not infection; organism not
needed after toxin produced
– Toxin not destroyed by proteases of digestive
tract; probably complexed with other proteins
? Mechanism
– Affects peripheral nerve endings
Lecture 9 BIOL 533 43
Botulinum Toxin
– Once across the gut,it is carried in the blood
to neuromuscular junctions
? Bind to gangliosides at motor nerve endpoints and
is taken up by cell
? Subsequent events unknown
– Result in presynaptic block of release of acetylcholine
– Interruptions in nerve stimulation causes irreversible
relaxation of muscles—leads to respiratory arrest
Lecture 9 BIOL 533 44
Lecture 9
? Questions?
? Comments?
? Assignments..,
