“SYNOPSIS”

BACTERIAL TOXINS

NAME : SHASHI PAUL

REG.NO. : 11006142

ROLL.NO : RP8003-B-15

COURSE : MSc.MICROBIOLOGY

COURSE CODE : 2403

SUBJECT CODE : BTY503

Submitted to:
Dr. Anjuman Sir.
ACKNOWLEGMENT

It has been a great challenge but a plenty of learning and

opportunities to gain huge knowledge on the way preparing this term
paper. I would not succeed without my teacher Dr.Anjuman , who
seemed to be with me always; and prepared to give me feedback and
guidelines whenever I needed it.

Thank You
Sir!
I also would like to thank all my friends.

I hope you will find my working as interesting and knowledge

earning. And it will be useful for others wanting to learn about apparel
industry and retailers’ policies and strategies about planning.

SHASHI PAUL................
 CONTENTS

 INTRODUCTION
 GENERAL ASPECT
 TOXIN TYPES
 TOXIN PRODUCTION
 MECHANISM OF ACTION
 DIPHTHERIA TOXINS
 CHOLERA TOXINS
 TETANUS TOXINS
 BOTULINUM TOXINS
 BACTERIAL TOXINS

HISTORY
Since diphtheria toxin was isolated by Roux and Yersin in 1888 (1),
microbial toxins have been recognized as the primary virulence factor(s) for
a variety of pathogenic bacteria. Bacterial toxins have been defined as
"soluble substances that alter the normal metabolism of host cells with
deleterious effects on the host"

INTRODUCTION
Bacterial toxins are by-products produced by pathogenic microbes that
have taken up residence in the body. Bacterium can enter a host by various
means, such as consuming contaminated food or water. Bacteria can also
be introduced through mucous membranes, either by direct contact with the
source or as a consequence of breathing in air-borne bacteria. The type of
bacterial toxins released depends on the species of invading bacteria.

GENERAL ASPECT
Many bacterial exotoxins have the capacity to damage the extracellular
matrix or the plasma membrane of eukaryotic cells. The damage not only
may result in the direct lysis of cells but also can facilitate bacterial spread
through tissues. Toxins that mediate this cellular damage do so by either
nzymatic hydrolysis or pore formation.
TOXIN TYPES

 Exotoxin
 Endotoxin

SPECIFICITY
- Some act on certain cell types
– Other affect wide range of cells and tissues

TOXIN PRODUCTION
 Found on phage
 Found on plasmids

MECHANISM OF ACTION
 Sphere of influence
 Level of toxicity
 Mechanisms of damage

TOXINS THAT ASSIST BACTERIAL SPREAD IN TISSUES

PROPERTIES
 Do not target any type of cell
 Include degradative enzymes that allow spreading
EXAMPLES
 Streptococci
TOXINS THAT LYSE CELLS

GENERAL ASPECTS
 Large class kill host cells by destroying their membranes; act as
lipases .
 Also hemolysins are of this type; lyse both red blood cells and
white blood cells .

MECHANISM
make membrane more permeable, water pours into cytoplasm, cell begins
to swell, and eventually bursts

EXAMPLES
 Streptococcal streptolysin O (heterogeneous pore former)

TOXINS THAT BLOCK PROTIEN 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

SIMILARTIES
 Most have two portions (A-B toxins)
Subunits

 Toxic activity (A)

 Binding to cell membrane (B)

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

DIPHTHERIA TOXINS
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

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+
EF2 is only known substrate for diphtheria toxin
  EF2 contains rare modification of one of histidine residues and
this is site recognized by toxin

MECHANISM OF ACTION
Pharmacological toxins
 Excess of cAMP interferes with phagocyte
functioning (chemotaxis and phagocytosis)

METHOD OF INCREASING

 Secretion of Camp

 Secretion of adenyl cyclase to make more cAMP

 Secretion of toxin alters activity of host adenyl cyclase

(cholera)

CHOLERA TOXINS
Target tissue is small intestine epithelium.
 Structure and mechanism of toxin
 Structure of subunits

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.
 A1 fragment is enzymatically active
REGULATION
 Normal
 Abnormal (cholera) normal action of R protein mimicked
by cholera toxin

MECHANISM OF ACTION
 Other toxins that activate adenylate cyclase:
 Toxins that block nerve function:
 General mechanism of both:

TETANUS TOXINS
Acts at distance from central nervous system.

BOTULINUM TOXINS
GENERAL ASPECT
 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

TERM PAPER
 “Bacterial Toxins”

HISTORY

Since diphtheria toxin was isolated by Roux and Yersin in 1888

(1), microbial toxins have been recognized as the primary virulence

factor(s) for a variety of pathogenic bacteria. Bacterial toxins have been
defined as "soluble substances that alter the normal metabolism of host
cells with deleterious effects on the host"

(2). Indeed, the major symptoms associated with disease caused by

Corynebacterium diphtheriae (diphtheria), Bordetella pertussis (whooping
cough), Vibrio cholerae (cholera), Bacillus anthracis (anthrax), Clostridium
botulinum (botulism), Clostridium tetani (tetanus), and enterohemorrhagic
Escherichia coli (bloody diarrhea and hemolytic uremic syndrome) are all
related to the activities of the toxins produced by these organisms. With the
recognition of the central role of toxin in these and other diseases has
come the application of inactive toxins (toxoids) as vaccines. Such toxoid
vaccines have had an important positive impact on public health.

INTRODUCTION
Bacterial toxins are by-products produced by pathogenic microbes that
have taken up residence in the body. Bacterium can enter a host by various
means, such as consuming contaminated food or water. Bacteria can also
be introduced through mucous membranes, either by direct contact with the
source or as a consequence of breathing in air-borne bacteria. The type of
bacterial toxins released depends on the species of invading bacteria.

The cellular structure of bacterium also influences what kinds of bacterial

toxins are produced. While all bacteria have single cells, there is a
difference between their outer membranes that results in two classifications
of bacteria: Gram-positive or Gram-negative. This distinction is visible when
subjected to a “Gram stain,” which is an injection of a purple dye and a
subsequent alcohol wash. Cells that retain the dye color are Gram-positive;
those that do not are Gram-negative.

There are several types of bacterial toxins that may infect the human body
at different sites. For instance, enterotoxins are toxic proteins generated in
the intestines. Neurotoxins specifically target nerve cells. In addition,
certain enzymes may be produced that can impair metabolic functioning.
However, there are two primary groups of bacterial toxins that the above
generally fall into in terms of mechanism: exotoxins and endotoxins.

Both Gram-positive and Gram-negative bacteria produce exotoxins, some

of which are quite poisonous. For example, tetanus is caused by a bacterial
toxin produced by Clostridium tetani that acts as a neurotoxin. Generally,
the severity of symptoms and rate of recovery depends on how the
infection occurs. However, it has been established that only a small amount
of the pure toxin will prove fatal. Fortunately, this bacteria, as well as other
exotoxins, can be adapted to produce preventative vaccines.

Endotoxins are released by Gram-negative bacteria. At first, they are not as

aggressively toxic as exotoxins due to the fact they remain largely
contained in the cellular walls of bacteria. However, as these cells complete
their life cycle and die, the circulating volume of this toxin increases. In
addition, they cannot be used to make vaccines.

Normally, the body attempts to eliminate bacterial toxins before they can
cause harm. The immune system is the first line of defense, but it may
become overwhelmed by the rate of bacterial replication. In fact,
inflammation is an indication that bacterial overgrowth is occurring. In this
case, the immune system will do the next best thing — move the bacteria
out of the way. Usually, fat cells are the selected storage sites, which can
lead to the formation of cysts and tumors.

The bacterial toxins are a major cause of diseases since they are
responsible for the majority of symptoms and lesions during infection
[Böhnel and Gessler, 2005; Blackall and Marques, 2004]. They can be
classified into two categories; (i) exotoxins, a soluble substance secreted
by bacteria in the host tissues, and (ii) endotoxins, generally residing
within the cell wall and released into host tissues upon cell death
[Prescott et al., 1993]. The exotoxins act at a distance from the site of
infection and can diffuse through the organism. The elucidation of the
cellular mechanism of action of the bacterial exotoxins remains a complex
problem, but they appear to share a common mechanism of action such
as (i) binding to specific receptors on the plasma membranes of the
sensitive cells, (ii) pore-formation, (iii) internalization or translocation
across the membrane barrier and (iv) direct secretion [Middlebrook and
Dorland, 1984; Popoff, 2005].
The exotoxins have a special affinity for particular tissues and may be
divided into three categories on the basis of the site affected: i)
neurotoxins act on nervous system, ii) enterotoxins on intestinal
mucosa, and iii) cytotoxins on general tissue [Prescott et al., 1993].
The neurotoxins recognize specific receptors on the unmyelinated
areas of the presynaptic membrane and inhibit acetylcholine release.
The enterotoxins act by activating adenylate cyclase or guanylate
cyclase [Fishman, 1990]. Some staphylococcal enterotoxins cause the
food poisoning syndrome [Dinges et al., 2000]. The cytotoxins act on
general tissues; for example, vacuolating cytotoxin is one of the most
important virulence factors produced by Helicobactor pylori, a
causative agent of severe gastric diseases such as ulcers and cancers
[Montecucco et al., 1999].
The bacterial toxins have a wide range of applications today. For
example the cholera toxin (CT) and the heat-labile toxin from E. coli
(LT) have been used as strong mucosal adjuvants in experimental
models [Bagley et al., 2002]; bacterial pore-forming hemolysins, such
as listeriolysin O, have the potential for use in cytosolic drug delivery
systems [Provoda and Lee, 2000]; botulinum toxin has been employed
in orthopedics, physiatrics, gastroenterology, gynecology, neurology,
pediatrics, general surgery, plastic surgery and several other
specialties and also to treat hyperhidrosis and wrinkles in dermatology
[Tamura and Chang, 2003]. The ability of the toxoid vaccine to induce
toxin-neutralizing antibodies has provided the basis for the use of
therapeutic antitoxins and immunoglobulins for the prophylaxis and
treatment of diseases caused by bacterial toxins. In this study, a
systematic attempt has been made to develop a method for predicting
bacterial toxins, their class (exotoxin or endotoxin) and sub classes of
exotoxins.
GENERAL ASPECT
Many bacterial exotoxins have the capacity to damage the extracellular
matrix or the plasma membrane of eukaryotic cells. The damage not only
may result in the direct lysis of cells but also can facilitate bacterial spread
through tissues. Toxins that mediate this cellular damage do so by either
enzymatic hydrolysis or pore formation. Bacterial hyaluronidases,
collagenases, and phospholipases have the capacity to degrade cellular
membranes or matrices. Specific examples of these types of toxins include
the -toxin of Clostridium perfringens, which has phospholipase C activity;
Streptococcus pyogenes streptokinase, which can hydrolyze plasminogen
to plasmin and dissolve clots; and the clostridial collagenases . Pore-
forming toxins, as the name suggests, disrupt the selective influx and efflux
of ions across the plasma membrane by inserting a transmembrane pore.

Bacterial toxins can also target and alter the function of a variety of cellular
proteins without directly killing the intoxicated cell. Toxin activation or
modification of secondary messengers can cause dramatic alterations to
signal transduction pathways critical in maintaining a variety of cellular
functions. To demonstrate the diversity among the toxins that belong to this
category, we will describe CNF type 1 and the heat-stable enterotoxins.

BACTERIAL SUPERANTIGENS: TOO MUCH OF A GOOD

THING
Several bacterial toxins can act directly on the T cells and antigen-
presenting cells of the immune system. Impairment of the immunologic
functions of these cells by toxin can lead to human disease. One large
family of toxins in this category are the pyrogenic toxin superantigens
(PTSAgs), whose hallmark biological activities include potent stimulation of
the immune cell system, pyrogenicity, and enhancement of endotoxin
shock (49-51). These stable, secreted toxins of 22 kDa to 30 kDa include
staphylococcal enterotoxins serotypes A-E, G, and H; group A
streptococcal pyrogenic exotoxins serotypes A-C and F; group A
streptococcal superantigen; and staphylococcal TSST-1, which we discuss
below.

DR. JEKYLL OR MR. HYDE ?

Some of these powerful disease-causing toxins have been exploited to
further basic knowledge of cell biology or for medical purposes. For
example, cholera toxin and the related labile-toxin of E. coli, as well as B.
pertussis toxin, have been used as biologic tools to understand the
mechanism of adenylate cyclase activation and the role of cyclic AMP as a
second messenger in the eukaryotic cell. Derivatives of some of these
toxins, cholera toxin and labile toxin, have also been incorporated into
human vaccines because of the adjuvant properties of these molecules.

Similarly, the activities of several potent cytotoxins have been harnessed

as potential therapies for certain cancers. Such toxins can either be used
directly in treatment or as components of immunotoxins . For example, Stx
binds to the cell surface glycolipid CD77, which is expressed by B cells in
certain B-cell lymphomas. This finding led to studies that showed that Stx
can purge murine (and potentially human) bone marrow of malignant
CD77+ B cells before an autologous bone marrow transplant . Other toxins
that inhibit protein synthesis, such as diphtheria toxin, Pseudomonas
exotoxin A, or the plant toxin ricin, are frequently engineered as the cell-
killing component of immunotoxins. These "magic bullets," hybrids of the
enzymatically active portion of a toxin molecule and monoclonal antibodies
(or a receptor), are in clinical trials for the treatment of persons with B-cell
lymphomas, leukemia, and bone marrow transplants.

Several clinical applications have also been found for the powerful
botulinum neurotoxin type A (BoNT/A). The disorders that respond to
BoNT/A involve muscle hyperactivity. A minuscule amount of purified toxin
injected into specific sites results in paralysis of the target muscle and
ablation of the muscle spasm. Therapy must be continual since the effect of
the toxin usually lasts for no more than several months. The first maladies
treated with BoNT/A were eye movement abnormalities. However, the
therapeutic value of BoNT/A has been shown for many other disorders
including cervical and laryngeal dystonia, writer's cramp, hemifacial spasm,
tremors, and tics. BoNT/A is also used cosmetically to reduce deep
wrinkles caused by the contraction of facial muscles.

Another toxic bacterial product with medical applications is streptokinase, a

potent plasminogen activator produced by several pathogenic streptococcal
strains. The proteolytic activity of streptokinase is used to clear blocked
arteries in patients who have heart attacks .

VACCINATE, DON’T PROCRASTINATE

Vaccines directed at the toxic component of bacterial pathogens have
proven quite effective in preventing certain diseases. Most licensed toxoid
vaccines are relatively crude, but effective, preparations. These vaccines
consist of partially purified toxin preparations obtained from culture
supernatants of bacteria such as C. diphtheriae, C. tetani, or B. anthracis.
Formaldehyde treatment is used to detoxify the diphtheria and tetanus
toxins for vaccine formulation. The anthrax vaccine contains the protective
antigen and small amounts of the lethal factor and edema factor toxins. The
current botulinum vaccine is an investigational drug composed of crude
preparations of five botulinum toxoids and is distributed by the Centers for
Disease Control and Prevention to researchers that work with the toxin or
organism. Acellular pertussis vaccines that contain pertussis toxoid, alone
or as one of several components, are as effective as killed whole-cell
vaccines but less reactogenic ; such vaccines have recently been approved
for use in infants as well as older children.

New vaccines aimed at toxins are in various stages of development:

research and development, preclinical, phase I, phase II, or phase III. The
next generation of toxoid vaccines falls into three general categories:
purified toxoids that have been inactivated by chemical or genetic means;
live, attenuated strains of the causative agent that produce a genetically
derived toxoid; or live, attenuated unrelated bacterial vector strains, such
as V. cholerae or Salmonella, that produce the target toxoid. Examples of
each of these approaches and progress in development of specific toxoid
vaccines are described annually in the Jordan Report.

Antitoxins raised against diphtheria, tetanus, and botulinum toxoids have

also been used for many years to treat seriously ill patients. Antiserum
specific for the Stx toxins produced by E. coli O157:H7 and other STEC is
under development for the treatment and prevention of hemolytic uremic
syndrome, a life-threatening sequela of these infections.

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

TOXIN TYPE

 EXOTOXINS
Protein produced by bacteria either excreted or bound to bacterial surface
and released when lysed

 ENDOTOXINS
Lps of the outer membrane of Gram-Bacteria acts as toxin only under
special circumstances

SPECIFICITY
 Some act on certain cell types
 Other affect wide range of cells and tissues
NUMBERS PRODUCED BY SINGLE BACTERIUM
 Some produce none
Pneumococci

TOXIN PRODUCTION
PROPERTIES
 Dispensable, but essential under certain situations where survival
and spread are at stake
 Genes frequently carried on plasmids and temperate
bacteriophage

FOUND ON PHAGE

toxin genes for:

 Diphtheria
 Botulism
 Scarlet fever
 Toxic streptococci (“flesh-eating”)

FOUND ON PLASMIDS

 E. colitoxin causes diarrhea

 S. aureustoxin causes “scalded skin syndrome”

 E. coli0157:H7
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)

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?

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

MECHANISM OF ACTION
SPHERE OF INFLUENCE
 Some act locally, killing WBC nearby
 Others help organism to spread in host tissues 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

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 diphtheria
- 1000-fold more for Pseudomonas A
MECHANISM 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

TOXINS THAT ASSIST BACTERIAL SPREAD IN TISSUES

PROPERTIES
 Do not target any type of cell
 Include degradative enzymes that allow spreading
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

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)

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
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

EXAMPLES
 Staphylococciα-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

EXAMPLES
 Streptococcal streptolysin O (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

TOXINS THAT BLOCK PROTIEN 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

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 internalization of the toxin (some investigators propose
direct passage through pore)
“A” moiety is often latent, even after engulfment
 May be activated by proteolytic cleavage and reduction of disulfide
bridges
 Toxins of diphtheria, cholera, tetanus, andS hige lla are synthesized as
inactive precursors
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)

DIPHTHERIATOXINS

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
 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

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+
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. aeruginosaexotoxin A works same as diphtheria toxin

MECHANISM OF ACTION
Pharmacological 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)

CHOLERA TOXINS
Target tissue is small intestine epithelium

STRUCTURE AND MECHANISM OF ACTION

  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

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)

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 is enzymatically active
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
 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

ABNORMAL (CHOLERA) NORMAL ACTION OFR PROTIEN

MIMICKED BY CHOLERA TOXINS
 Gs remains in active state to stimulate adenyl cyclase
 Promotes active state of Gs protein by different mechanism
  ADP-ribosylates Gs at one of its arginine residues (Gs protein
locked into active conformation)

MECHANISM OF ACTION
Other toxins that activate adenylate cyclase
 Number of enterotoxins that produce diarrhea LT (labile)E.coli
 Bordetella pertussisadenylate cyclase.
Raise level cAMP in leucocytes
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
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

TETANUS TOXINS
Acts at distance from central nervous system.
  Once bound to cell membranes, toxin is internalized probably
by receptor-mediated endocytosis
 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

BOTULINUM TOXINS

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
 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.

SUMMARY
Microbial toxins capable of interrupting or hyperstimulating many essential
functions and pathways of eukaryotic cells have evolved along with the
carrier bacterium. Presumably these toxins confer some benefit to the
bacterium, either during a stage of the host-parasite interaction or in some
environmental niche encountered by the bacterium. Certain bacterial toxins
act on the target cell surface to irreparably damage the cell membrane or
alter normal cellular signal transduction. Other toxins exhibit enzymatic
activity once the molecule has gained access to the cytoplasm of the
sensitive cell by endocytosis. Yet other bacterial toxins act by either turning
off or locking on a normal host cell function.

Although detrimental to the susceptible host during an infection, the

activities of several bacterial toxins have been exploited as probes of
eukaryotic cellular pathways and for medicinal applications. Thus, research
on a microbial toxin produced by an established, emerging, or reemerging
pathogen is likely to yield novel information about the role of that toxin in
disease as well as the properties of host cells that are subverted by the
toxin.

REFERENCE
1. WIKIPEDIA.COM
2. http://en.wikipedia.org/wiki/Template:Phospholipids
3. Roux E, Yersin A. Contribution a l'etude de la diphtherie. Annales de l'Institut Pasteur
1888;2:629-61.
4. Schlessinger D, Schaechter M. Bacterial toxins. In: Schaechter M, Medoff G, Eisenstein
BI, editors. Mechanisms of microbial disease. 2nd ed. Baltimore: Williams and Wilkins;
1993. p. 162-75.
5. Songer JG. Bacterial phospholipases and their role in virulence. Trends Microbiol
1997;5:156-61.
6. Lottenberg R, Minning-Wenz D, Boyle MD. Capturing host plasmin(ogen): a common
mechanism for invasive pathogens? Trends Microbiol 1994;2:20-4.
7. Harrington DJ. Bacterial collagenases and collagen-degrading enzymes and their
potential role in human disease. Infect Immun 1996;64:1885-91.
8. Bhakdi S, Tranum-Jensen J. Alpha-toxin of Staphylococcus aureus. Microbiological
Reviews 1991;55:733-51.
9. Tomita T, Kamio Y. Molecular biology of the pore-forming cytolysins from
Staphylococcus aureus, a- and gamma-hemolysins and leukocidin. Bioscience,
Biotechnology, and Biochemistry 1997;61:565-72.
10. Bhakdi S, Bayley H, Valeva A, Walev I, Walker B, Weller U, et al. Staphylococcal
alpha-toxin, streptolysin-O and Escherichia coli hemolysin: prototypes of pore-forming
bacterial cytolysins. Arch Microbiol 1996;165:73-9.
11. Song L, Hobaugh MR, Shustak C, Cheley S, Bayley H, Gouaux JE. Structure of
staphylococcal alpha-hemolysin, a heptameric transmembrane pore. Science
1996;274:1859-66.
12. Lesieur C, Vecsey-Semjen B, Abrami L, Fivaz M, Gisou van der Goot F. Membrane
insertion: the strategies of toxins. Mol Membr Biol 1997;14:45-64.
13. Collier RJ. In: Moss J, Vaughan M, editors. ADP-ribosylating toxins and g proteins.
Washington: American Society for Microbiology; 1990. p. 3-19.
14. Wick MJ, Iglewski BH. In: Moss J, Vaughan M, editors. ADP-ribosylating toxins and g
proteins. Washington: American Society for Microbiology; 1990. p. 31-43.
15. Endo Y, Tsurugi K, Yutsudo T, Takeda Y, Ogasawara Y, Igarashi K. Site of action of a
Vero toxin (VT2) from Escherichia coli O157:H7 and of Shiga toxin on eucaryotic
ribosomes. Eur J Biochem 1988;171:45-50.
16. Saxena SK, O'Brien AD, Ackerman EJ. Shiga toxin, Shiga-like toxin II variant, and ricin
are all single-site RNA N-glycosidases of 28 S RNA when microinjected into Xenopus
oocytes. J Biol Chem 1989;264:596-601.
17. Tesh VL, O'Brien AD. The pathogenic mechanisms of Shiga toxin and the Shiga-like
toxins. Mol Microbiol 1991;5:1817-22.
18. O'Brien AD, Tesh VL, Donohue-Rolfe A, Jackson MP, Olsnes S, Sandvig K, et al. Shiga
toxin: biochemistry, genetics, mode of action, and role in pathogenesis. In: Sansonetti PJ,
editor. Pathogenesis of shigellosis. 180th ed. Berlin-Heidelberg: Springer-Verlag; 1992.
p. 66-94.
19. O'Brien AD, Kaper JB. Shiga toxin-producing Escherichia coli: yesterday, today, and
tomorrow. In: Kaper JB, O'Brien AD, editors. Escherichia coli O157:H7 and other Shiga
toxin-producing E. coli strains. Washington: American Society for Microbiology; 1998.
p. 1-11.
20. Melton-Celsa AR, O'Brien AD. Activation of Shiga-like toxins by mouse and human
intestinal mucus correlates with virulence of enterohemorrhagic Escherichia coli
O91:H21 isolates in orally infected, streptomycin-treated mice. Infect Immun
1996;64:1569-76.
21. Stein PE, Boodhoo A, Tyrell GT, Brunton J, Read RJ. Crystal structure of the cell-
binding B oligomer of verotoxin-1 from E. coli. Nature 1992;355:748-50.
22. Frasier ME, Chernaia MM, Kozlov YV, James MNG. Crystal structure of the holotoxin
from Shigella dysenteriae at 2.5 Å resolution. Nature Structural Biology 1994;1:59-64.