MICROBIOLOGY LECTURE

CHAPTER 2 – MICROBIAL CELL STRUCTURE AND FUNCTION | CHAPTER 5 – VIRUS

THE CELL ENVELOPE BACTERIAL CYTOPLASMIC MEMBRANES

→ Cell envelope consists of a series of → Cytoplasmic membrane of all bacterial

layered structures.
• It surrounds the cytoplasm
• It govern cellular interaction with the
external environment.
→ Important functions of cell envelope:
• Governs transport of nutrients into the
cell and wastes out of the cell
• Site of energy conservation
• Governs cell shape
• Protects cell from mechanical stress
• Help the cell attach to surfaces
• Protect cell from attacks
THE CYTOPLASMIC MEMBRANE
and eukaryal cells is a phospholipid
bilayer containing embedded proteins.
→ Cytoplasmic membrane is 8-10
nanometers wide.
→ Phospholipids are composed of
hydrophobic (water-repelling) and
hydrophilic (waster-attracting)
components.
→ Hydrophobic component consist of fatty
acid “tails”
→ Hydrophilic component consists of
glycerophosphate and one of several
other functional groups also bonded to
the phosphate.

→ Cytoplasmic membrane: surrounds the → The membrane is bilayer:

cytoplasm and separates it from the
environment.
→ Cytoplasmic membrane is rather weak
(physically)
→ Has ideal structure for its major function:
selective permeability.
• Two phospholipid layers
• Fatty acid tails face each other
creating a hydrophobic region.
• Hydrophilic heads are exposed to the
environment.

→ Transportation of nutrients inwards and

waste products outwards occur across
the cytoplasmic membrane.

→ Proteins located in the cytoplasmic

membrane facilitate these reactions.
• Many other membrane proteins play
important roles in energy metabolism.
→ Variety of proteins are integrated into the • In Archaea: the lipids have
cytoplasmic membrane. hydrophobic isoprenoid bound to
• Membrane proteins typically have glycerol by ether bonds.
hydrophobic domains and hydrophilic
domains. → Hydrophobic region of archaeal
membranes is formed from repeating
→ Proteins embedded in the membrane are units isoprene than fatty acids.
integral membranes
→ The cytoplasmic membrane of Archaea is
→ Many integral membrane proteins extend constructed from diphosphoglycerol
across the membrane and these are tetraethers.
called transmembrane protein • In tetraether lipid structure, the ends
of the inwardly pointing isoprenoid
→ Peripheral membrane proteins are more chains are covalently linked.
loosely attached. • Forms a lipid monolayer instead of
lipid bilayer.
• Some peripheral membrane proteins
are lipoproteins: contains a
hydrophobic lipid tail that anchors the
protein into the membrane.
• Others have residues that associate
with the hydrophilic head groups or
associate indirectly with membranes
by binding to other proteins.
• Typically interact with integral
membrane proteins in important
cellular processes.

→ Archaeal lipids can have many different

isoprenoid chains including some that
contain ring structures.
• Example: Crenarchaeol

→ Despite differences in chemistry between

Archaea and other organisms in the other
phylogenetic domains, the fundamental
construction is the same in all cells.

ARCHAEAL CYTOPLSMIC MEMBRANES CYTOPLASMIC MEMBRANE FUNCTION

→ Cytoplasmic membrane of Archaea is

structurally similar to Bacteria and
Eukarya, but chemistry is different.
• In Bacteria and Eukarya:
hydrophobic fatty acid tails are
bound to glycerol by ester linkages.
→ The cytoplasmic membrane has at least
three major functions:
1. Cell’s permeability barrier: prevents
passive leakage of solutes into or
out of the cell.
→ The cytoplasmic membrane is a barrier to
the diffusion of most substances.
→ Most substances that enter of leave the
cell must be carried by transport
proteins
• Because the cytoplasmic membrane
is impermeable
• Function accumulate solutes against
the concentration gradient
• Process that diffusion alone cannot
do.
→ Transport (requires energy) ensures that
the cytoplasm has sufficient concentration
of the nutrients it needs.
→ Transport proteins typically display high
sensitivity and high specificity.
→ Transport proteins transport only a single
kind of molecule while others carry a
related class of molecules.
• This economizing reduces the need
for separated transport proteins for
each different sugar or amino acid.
2. Anchors several proteins that
catalyze a suite of key cell
functions.
3. Cytoplasmic membrane of Bacteria
and Archaea plays a major role in
energy conservation and
consumption.
→ The cytoplasmic membrane can be
energized when protons are separated
from hydroxyl ions.
• Charge separation creates an
energized state called the proton
motive force.
• Dissipation of the proton motive force
can be coupled to several energy
requiring actions
• Examples: transport, cell locomotion,
biosynthesis of ATP
TRANSPORTING NUTRIENTS INTO THE
CELL
ACTIVE TRANSPORT AND
TRANSPORTERS
→ Active Transport: process by which cells
accumulate solutes against the
concentration gradient
→ Three basic mechanisms of active
transport:
1. Simple transport system consists
only of a transmembrane transport
protein.
2. Group translocation employs a
series of proteins in the transport
event.
3. ABC transport systems consist of a
binding protein, transmembrane
transporter and ATP hydrolyzing
protein
→ Each of these transport systems is
energy driven.
• Driven by proton motive force, ATP, or
other energy rich compound.
→ Transmembrane component of all
transport systems is composed of a
polypeptide containing 12 regions.
• Polypeptide weave back and forth
through the membrane to form a
channel
• Through this channel that the solute
is transported into the cell.
→ Transport is linked to conformational
change that occurs when it binds to a
specific solute.
 • Like a gate swing open, this
conformational change sweeps the
solute into the cell.
SIMPLE TRANSPORTER AND GROUP
TRANSLOCATION
→ Simple transport reactions are driven by
the energy inherent in the proton motive
force.
→ Two major transport events catalyzed are:
• Symport reactions: a solute and a
proton are cotransported in the same
direction
• Antiport reactions: solute a proton
are transported in opposite direction.
→ Example of simple transporter: uptake of
sugar lactose by way of the lac
permease.
• A well-studied symporter in
Escherichia coli.
• As each lactose molecule enters the
cell, the potential energy in the proton
motive force is diminished slightly by
the cotransport of a proton.
• The net result is the energy driven
accumulation of lactose in the
cytoplasm against the concentration
gradient.
→ Group translocation differs from simple
transport in two important ways:
• Transported substance is chemically
modified
• Energy rich organic compound drives
the transport event.
→ Example of group translocation:
• Transport the sugars glucose,
mannose, and fructose in E. coli
• During uptake, these compounds are
phosphorylated by the
phosphotransferase system.
• Energy to drive the
phosphotransferase system come
from phosphoenolpyruvate – an
energy rich intermediate in glycolysis.
ABC TRANSPORTER SYSTEMS
→ ABC Transporters are modular systems
that have three components:
• Binding protein
• Transmembrane protein channel
• ATP hydrolyzing protein
→ ABC stands for ATP binding cassette –
a structural feature of proteins that bind
ATP.
→ More than 200 different ABC transport
systems are known.
• These catalyze the uptake of a wide
variety of organic and inorganic
compounds.
→ Substrate-binding proteins are present
outside of the cell.
• They bind to a specific substrate and
enable transport into the cell.
→ A characteristic property of binding
proteins is their extremely high substrate
affinity.
• These proteins can bind their specific
substrate even when it is present at
extremely low concentration.
→ Once its specific substrate is bound, the
binding protein interacts with its
respective transmembrane component to
transport the substrate into the cell driven
by the energy in ATP
THE CELL WALL
→ Cytoplasm of prokaryotic cells maintains
a high concentration of dissolved solutes
that creates significant osmotic pressure.
 • The osmotic pressure is enough to
cause the cell membrane to burst and
the cell to die.
• The process is called cell lysis.
→ To withstand the turgor pressure, the cell
envelopes of most Bacteria and
Archaea have a layer outside the
cytoplasmic membrane called the cell
wall
→ Cell wall is for:
• Protection against osmotic lysis
• Maintains cell shape and rigidity
→ The cell envelopes of most Bacteria can
be classified as:
• Gram positive
• Gram negative
• This is based on their organization
and cell wall structure.
→ Gram positive cell: contains a cytoplasmic
membrane and a thick cell wall
→ Gram negative cell: cytoplasmic
membrane, thin cell wall, outer
membrane, periplasm.
• Periplasm is a compartment between
the cytoplasmic and outer
membranes.
→ Th gram stain reaction is determined
primarily by the thickness of the cell wall
rather than the number of layers in the
cell envelope.
• Gram stain reaction does not always
corelate with cell envelope structure.
→ The gram stain reaction is sufficiently
predictive of cell envelope structure in
Bacteria.
• The names of the two most common
bacterial cell envelopes (gram
negative and gram positive) are based
on their reactions to gram stains.
→ Knowledge of cell wall envelope structure
and function is important.
• Example: certain antibiotic like the
penicillins and cephalosporins target
bacterial cell wall synthesis, leaving
the cell susceptible to osmotic lysis.
• Since human and other animal cells
lack cell walls and are not a target of
such antibiotics, these drugs are for
treating bacterial infections.
BACTERIAL CELL WALLS
→ Cell wall found in bacteria contain a rigid
polysaccharide called peptidoglycan.
• It confers structural strength on the
cell.
→ Peptidoglycan is found in all Bacteria that
contain a cell wall.
• Only unique to Bacteria and not found
in Archaea and Eukarya
→ Sugar backbone of peptidoglycan is
composed of alternating repeats of two
modified glucose residues called N-
acetylglucosamine (G) and N-
acetylmuramic acid (M).
• Attached to the latter is a short
peptide side chain.
• Glycan tetrapeptide: L-alanine, D-
alanine, D-glutamic acid,
diaminopimelic acid (DAP)
• The peptide side chain can vary
between bacterial species
→ Strands of peptidoglycan run parallel to
each other around the circumference of
the cell.
 • The peptide side chains of adjacent
peptidoglycan strands are cross
linked by peptide bonds.
• Peptidoglycan the forms one single
molecule
→ The cell wall in gram negative cell
envelope is 2-7 nm thick
• Consists of a single layer of
peptidoglycan.
• Can be up to three layers.
→ The peptidoglycan mesh is flexible and
porous.
• But is strong enough to resist turgor
pressure and prevent rupture of
membrane and cell lysis.
→ Typical gram positive cell envelope
contains a thick peptidoglycan cell.
• Can measure up to 20-35 nm in
thickness.
• Much thicker than gram negative.
→ 90% of gram positive cell envelope can
consist of peptidoglycan.
→ Gram negative cell wall typically consists
of single layer
→ Gram positive cell wall can be 15 or more
layers thick.
→ Gram positive cell wall is stabilized three
dimensionally by peptide cross links,
which forms between adjacent
peptidoglycan strands (horizontally and
vertically)
→ Many gram positive bacteria produce
acidic molecules called teichoic acids
embedded in their cell wall
• Teichoic acids are composed of
glycerol phosphate or ribitol
phosphate with attached molecules of
glucose or D-alanine.
• Individual alcohol molecules are then
connected through the phosphate
groups to form long strands.
• These are then covalently linked to
peptidoglycan.
→ Lipoteichoic acids: teichoic acids are
covalently bonded to membrane lipids
rather than peptidoglycan.
→ Peptidoglycan can be destroyed by
lysozyme – an enzyme that cleaves the
glycosidic bond between N-
acetylglucosamine and N-acetylmuramic
acid
• This weakens the peptidoglycan and
can cause cell lysis.
→ Lysozyme is present in human secretions
including tears, saliva, and other bodily
fluids.
• Functions as a major line of defense
against bacterial infections.
→ Penicillin, also targets peptidoglycan.
• Penicillin blocks the formation of
peptide cross links, which
compromises the strength of
peptidoglycan, leading to cell lysis.
ARCHAEAL CELL WALLS
→ Archaea’s cell wall lacks peptidoglycan.
→ Archaea typically lack an outer
membrane.
→ Gram stain reaction is not useful for
predicting the structures of archaeal cell
envelopes.
• We do not use terms like gram
negative a positive for to describe
Archaea cells.
→ Most Archaea lack a polysaccharide-
containing cell wall (like peptidoglycan)
• Have an S layer instead
• S layer: rigid protein shell that
functions to prevent osmotic lysis
same with bacterial cell wall.
→ Some Archaea have cells walls
• Cell wall have unique chemical
structure not found in bacteria.
• Example: cell walls of methane-
producing Archaea contain a
polysaccharide called pseudomurein.
o Pseudomurein is structurally
similar to peptidoglycan
o Its backbone is formed from
alternating repeats of N-
acetylglucosamine and N-
acetyltalosaminuronic acid.
→ It is likely that peptidoglycan and
pseudomurein are variants of a cell wall
polysaccharide originally present in the
common ancestor of Bacteria and
Archaea.
→ Pseudomurein is immune from
destruction by both lysozyme and
penicillin.
LPS: THE OUTER MEMBRANE
→ Most of the gram negative cell envelope
is composed of the outer membrane.
→ Outer membrane: second lipid bilayer
found external to the cell wall.
• Structure and function differs from the
cytoplasmic membrane.
→ Outer membrane and cytoplasmic are
similar because they both contain
phospholipid and protein.
→ The outer membrane also contains
polysaccharide molecules covalently
bound to lipids.
• Outer membrane is often called the
liposaccharide layer or LPS
→ LPS molecules have several unique
functions
• Facilitate surface recognition
• Virulence factors for some bacterial
pathogens
• Contribute to mechanical strength of
cell wall
→ Outer membrane also contains porins –
transmembrane proteins that allow for
nonspecific transport of solutes.
STRUCTURE AND ACTIVITY OF LPS
→ LPS contains a polysaccharide that
consists of the core polysaccharide and
O-specific polysaccharide
→ The core polysaccharide consists of:
• Ketodeoxyoctonate (KDO)
• Seven carbon sugars (heptoses)
• Hexose sugars: glucose, galactose,
and N-acetylglucosamine.
→ The O-specific polysaccharide is
connected to the core and consist of:
 • Galactose
• Glucose
• Hexoses: rhamnose and mannose
• One or more dideoxyhexoses:
abequose, colitose, paratose or
tyvelose.
→ Within the outer membrane, these
negatively charged polysaccharides can
be linked together when adjacent LPS
molecules form ionic bonds.
• The ionic bonds confers strength to
the outer membranes.
→ The lipid portion of the LPS is called lipid
A • Some endotoxins cause violent
• Is not a typical glycerol lipid
• Fatty acids are bonded through the
amine groups from a disaccharide
composed of glucosamine phosphate.
• The disaccharide is attached to the
core polysaccharide through KDO
→ Fatty acids typically found in lipid A are:
• Caproic
• Lauric
• Myristic
• Palmitic
• Stearic
→ LPS replaces much of the phospholipid in
the outer half of the cytoplasmic
membrane
→ Outer membrane is anchored to the
peptidoglycan layer by the Braun
lipoprotein
• Braun lipoprotein: molecule that spans
the gap between the LPS layer and
the peptidoglycan layer.
→ Important biological activity of LPS is its
toxicity to animals.
→ Common gram negative pathogens for
humans include species of:
• Salmonella
• Shigella
• Escherichia
→ Some of the gastrointestinal symptoms of
the pathogens elicit are due to their toxic
outer membrane components
→ Toxicity is linked to the LPS layer
specifically to lipid A
→ The term endotoxin refers to this toxic
component of LPS
symptoms to humans like:
o Gas
o Diarrhea
o Vomiting
→ The endotoxins produced by Salmonella
and enteropathogenic strains of
Escherichia coli are transmitted in
contaminated foods are examples.
THE PERIPLASM AND PORINS
→ The outer membrane is impermeable to
proteins and other large molecules
→ Another major function of the outer
membrane:
• Prevention of cellular proteins whose
activities must occur outside the
cytoplasm from diffusing away from
the cell
• These extracellular proteins reside in
the periplasm
→ Periplasm: a space of about 15 nm
located between the outer surface of the
cytoplasmic membrane and the inner
surface of the outer membrane
→ The periplasm may contain several
classes of proteins including:
 • Hydrolytic enzymes – function in the
initial degradation of polymeric
substances
• Binding proteins – begins the
process of transporting substances
• Chemoreceptors – govern the
chemotaxis response
• Proteins that construct
extracellular structures from
precursor molecules
→ Most periplasmic proteins reach the
periplasm by way of a protein-exporting
system present in the cytoplasmic
membrane
→ Outer membrane is relatively permeable
to small molecules because of proteins
called porins.
→ Porins: channels for the entrance and
exit of solutes
• Unique to the outer membrane of
Bacteria
• Should not be confused with
aquaporins
• Porins ≠ Aquaporins
→ Nonspecific porins form water-filled
channels through which most very small
hydrophilic substances can pass
→ Specific porins contain a binding site for
one or a group of structurally related
substance
→ Porins are transmembrane proteins
composed of three identical polypeptides
• The proteins are arranged to form
channels through which solutes can
diffuse
DIVERSITY OF CELL ENVELOPE
STRUCTURE
One way in which cell envelopes can vary
between cells is in the presence of an S-layer.
S-LAYERS
→ S-layers are found in many Bacteria and
in nearly all Archaea.
→ S-layer consists of a paracrystalline
monolayer of interlocking molecules of
protein or glycoprotein
→ S-layer is always the outermost layer of
the cell envelope.
→ S-layer is composed of only one or a few
subunits organized into repeating
structures.
• Can have hexagonal, tetragonal, or
trimeric symmetry.
• The repeating units form a rigid yet
permeable paracrystalline lattice
• Can be thick as 5-20 nm thick in
Bacteria
• 70 nm thick in some Archaea.
→ In Archaea, thick S-layers can take on the
role of the cell wall. Responsible for
structural strength, protection from lysis,
and conferring cell shape.
• Can also create a periplasmic like
space.
→ S-layers function as molecular sieves and
have pore sizes in the range of 2-10 nm
in diameter.
• The pores are large enough to allow
low-molecular weight compounds to
pass but small enough to trap large
molecules.
→ The compartment formed between the
cytoplasmic membrane and S-layer
functions like a periplasm.
→ As the outermost layer, S-layers can also
facilitate cell surface interactions such as
attachment
• Can also increase the ability of some
bacterial pathogens to cause disease
by either promoting adhesion or
protecting the cell from host
defenses.
ALTERNATIVE CONFIGURATIONS OF THE
CELL ENVELOPE
→ Most common components found in the
cell envelope:
• Cytoplasmic membrane (CM)
• Cell wall (CW)
• Outer membrane (OM)
• S-layer (SL)
→ A common variation on cell envelope
structure is to find an outer S-layer
surrounding an otherwise gram-positive
or negative bacterium.
→ Many Archaea have only an S-layer
outside of the cytoplasmic membrane.
• These layers are usually constructed
with paracrystalline or glycoprotein but
can still vary in their molecular
structures.
→ Some methanogenic Archaea have cell
walls made of pseudomurein.
• Will tend to may or may not have an
outer S-layer.
→ Archaea such as the heat-loving
Ignicoccus actually have an outer
membrane.
• Structure in Ignicoccus is unlike that
of gram negative bacteria
• It is composed largely of archaeal
isoprenoid lipids and lacks LPS
→ Although uncommon, a few Bacteria and
Archaea cell walls altogether.
• These include the mycoplasmas and
other pathogenic Bacteria that grow
within a host cell, and Archaea such
as Thermoplasma.
→ Lacking a cell wall, cells would be
expected to contain unusually tough
cytoplasmic membranes.
→ Example: Mycoplasmas
• most mycoplasmas contain sterols in
their cytoplasmic membranes.
o The sterols function to add
strength and rigidity.
• Mycoplasmas may also have little
need for a cell wall because they
experience little osmotic pressure
when living in the cytoplasmic
membrane of another cell
• The loss of peptidoglycan may help
mycoplasmas evade the host immune
system
o Host defenses recognize
bacterial cell wall components as
a signal of bacterial invasion.
CELL SURFACE STRUCTURE AND
INCLUSIONS
CELL SURFACE STRUCTURES
→ Many Bacteria and Archaea secrete
sticky or slimy materials on their cell
surface that consist of either
polysaccharide or proteins.
• The terms “capsule” and “slime layer”
are used to describe these layers.
→ These outer layers can:
• Meditate attachment
 • Protect cell from attack and
environmental stress
• Alter the diffusive environment of the
cell.
CAPSULES AND SLIME LAYERS
→ Capsule and slime layer are used to
describe a sticky coat of polysaccharide
formed outside of the cell envelope.
→ Capsule: polysaccharide layer is
organized in a tight matrix that excludes
small particles and is tightly attached to
the cell.
→ Capsules are readily visible by light
microscopy if cells are treated with India
ink.
• India ink contains particulates that
stains the background but cannot
penetrate the capsule
→ Slime layer: surface layer is easily
deformed and loosely attached, it will not
exclude particles and is more difficult to
see microscopically.
• Has a loosely attached polysaccharide
coat
• Easily detected in colonies of slime
forming species such as the lactic acid
bacterium Leuconostoc.
→ Surface polysaccharides assist in the
attachment of microorganisms to solid
surfaces.
• Pathogenic microorganisms that
enter the body usually do so by first
binding to specific surface
components of host tissues.
• Binding is often facilitated by bacterial
cell surface polysaccharides.
→ Many bacteria will bind to solid surfaces,
forming a thick layer of cells called
biofilm.
• Extracellular polysaccharides play a
key role in the development and
maintenance of biofilms
→ Outer surface layers also contribute to the
infectivity of a bacterial pathogen and
preventing dehydration.
• Example: the causative agents of
anthrax and bacterial pneumonia –
Bacillus anthracis and Streptococcus
pneumoniae each contains a thick
capsule of protein or polysaccharide
• Encapsulated cells of these bacteria
avoid destruction by the hosts
immune system because the immune
cells are blocked by the bacterial
capsule.
• Bacterial outer surface layers bind
water and this helps protect the cell
from dryness.
FIMBRIAE, PILI, AND HAMMI
→ Pili are thin (2-10nn) filamentous
structures made of protein that extend
from the surface of a cell.
• Short pili that mediate attachment are
often called fimbriae.
→ Pili enable bacterial cells to stick to
surfaces, including animal tissues, or to
form pellicles, or biofilms on solid
surfaces.
→ All gram negative bacteria produce pili
and many positive bacteria contain these
structures.
→ Pili contribute to the virulence of → The SM1 group inhabits anoxic
pathogens by allowing bacteria to attach groundwater in Earth’s deep subsurface
to other cells. • Hami function to affix cells to a
surface to form a networked biofilm.
→ Pili are diverse and can have other
important functions as well. → Hami structurally resemble type IV pili
except for their barbed terminus, which
→ Conjugative pili: facilitate genetic functions to attach cells to both surfaces
exchange causing cell to cell attachment and to each other.
during conjugation.
→ The biofilms formed by SM1 Archaea are
→ Electrically conductive pili: can conduct likely an ecological strategy that allows
electrons toward or away from the cell. these microbes to more efficiently trap the
• Play an important role in the energy scarce nutrient present in their habitat.
metabolism of diverse microbes.
→ Although cells of SM1 group are not as
→ Type IV pili: facilitate adhesion and also small as the groundwater
support an unusual form of cell ultramicrobacterial cells, they are less
movement called “twitching motility” in than 1um
certain bacterial species. • Their hami likely play an important
• Twitching motility: allows cells to move role in preventing cells from being
along solid surface washed away.
• In twitching motility, pili are extended
away from the cell, attach to a
surface, and are subsequently
retracted, dragging the cell forward.
• ATP supplies the energy necessary
• On rod shaped cells that move by
twitching, type IV pili are present only
at the cell poles
• Type IV pili assist in infectivity by
certain pathogens, including the gram
negative bacteria Vibrio cholerae
(cholera) and Neisseria gonorrhoeae
(gonorrhea) and the gram positive
bacterium Streptococcus pyogenes.
• Twitching motility assists organisms in
locating specific sites for attachment
to initiate disease process
• Type IV pili are also widespread in CELL INCLUSIONS
Archaea for surface adhesion and cell
aggregation.
→ Prokaryotic cell often contain inclusions.
→ An unusual group of Archaea, the SM1 • Many inclusions store energy or
group forms a unique structure called a nutrients
hamus (hami) • Some have other highly specialized
• Hamus (plural hami): resembles a functions that confer unique
grappling hook. properties
→ Inclusions are often visible in cells with
the light microscope.
→ Inclusions are enclosed by a single layer
membrane composed of proteins that
partitions off the inclusion in the
cytoplasm
• Like PHA, glycogen is a reservoir of
both carbon and energy and is
produced when carbon is in excess.
→ Glycogen resembles starch but differs in
the manner in which the glucose units are
linked together.

→ Storing carbon or other substances in an

insoluble form within the cytoplasm
reduces osmotic stress and takes up less
space compared with storing these
substances in soluble form.

CARBON STORAGE POLYMERS

→ Poly-β-hydroxybutyric acid (PHB):

most common inclusion bodies in
prokaryotic organisms.
• It is a lipid that is formed from β-
hydroxybutyric acid units.

→ Monomers of PHB polymerize by ester

linkage and then the polymer aggregates
into granules.
• The granules can be seen by light
and electron microscopy.

→ The monomer in polymer is usually

hydroxybutyrate (C4) but can vary in
length from as short as C3 to as long as
C18
PHOLYPHOSPHATE, SULFUR, AND
→ The more generic term poly-β-
CARBONATE MINERALS
hydroxyalkanoate (PHA) is often used
to describe this class of carbon and
energy storage polymers
→ Many prokaryotic and eukaryotic
microbes accumulate inorganic
→ PHAs are synthesized by cells when phosphate (PO43-) in the form of
there is an excess of carbon and are polyphosphate granules.
broken down as carbon or energy • Granules are formed when phosphate
sources when conditions warrant. is in excess and can be drawn upon
as a source of phosphate
→ Another carbon storage inclusion is • Source for nucleic acid and
glycogen phospholipid biosynthesis when
phosphate is limiting.
→ Glycogen: is a polymer of glucose
 • In some organisms, polyphosphate
can be broken down to synthesize
ATP from ADP
→ Many gram-negative Bacteria and
Archaea oxidize reduced sulfur
compounds such as hydrogen sulfide.
• These organisms are the “sulfur
bacteria” discovered by Sergei
Winogradsky
→ The oxidation of sulfide generates
electrons for use in energy metabolisms
or CO2 fixation
→ Elemental sulfur from oxidation of sulfide
may accumulate in the cell in
microscopically visible granules.
• This sulfur remains as long as the
source of reduced sulfur is still
present
• As the reduced sulfur source
becomes limiting, the S0 in the
granules is oxidized to sulfate and the
granules disappear
• Sulfur globules are present in the
periplasm.
• The periplasm expands outward to
accommodate the growing globules
as H2S is oxidized to S0 and then
contracts inward as S0 is oxidized to
SO42-
→ Some cyanobacteria form carbonate
minerals inside the cell, as cell inclusions
→ Example: Gloeomargarita forms
intracellular granules of benstonite
• Benstonite: carbonate mineral that
contains barium, strontium, and
magnesium.
• Benstonite might function as ballast to
maintain cell of this cyanobacterium in
their habitat
• The mineral could also be a way to
sequester carbonate to support
autotrophic growth.
→ Biomineralization: microbiological process
of forming minerals
GAS VESICLES
→ Some Bacteria and Archaea can float
because the contain gas vesicles.
→ Gas vesicles: structures that confer
buoyancy and allow the cells to position
themselves in regions of the water
column that best suit their metabolisms.
→ Example: cyanobacteria that form
massive accumulations called blooms in
lakes or other water bodies.
• Blooms are on or near the lake
surface where sunlight is most intense
and photosynthesis can occur at
maximal rates.
→ Gas vesicles are conical shaped
structures composed of two different
proteins
• Are hollow yet rigid and varies in
length and diameter.
→ Gas vesicles in different species vary
from 300-more than 1000 nm in length
and 45-120 in width.
→ Gas vesicles may number from a few to
hundreds per cell.
→ Impermeable to water and solutes.
→ Permeable to gas
• The composition and pressure of the
has inside a vesicle is that in which
the organism is suspended.
• Could be air at 1 atm in cyanobacteria
on a lake surface
• Could also be a mixture of gases as
N2, CO2, and H2 at greater than 1 atm
in species that inhabit anoxic zones
deeper in the lake.
→ Presence of gas vesicles can be detected
either by light microscopy.
 • Clusters of vesicles called gas
vacuoles, appear as irregular bright
inclusions
• Can also be seen by transmission
electron microscopy of cell thin
sections.
MAGNETOSOMES
→ Some bacteria can orient themselves
within a magnetic field because they
contain magnetosomes
→ These structure are biomineralized
particles of the magnetic iron oxides
magnetite [Fe(II)Fe(III)2O4] or greigite
[Fe(II)Fe(III)2S4]
→ Magnetosomes impart and magnetic
dipole on a cell.
• This allows it to orient itself in a
magnetic field
• Allows the cell to undergo
magnetotaxis – process of migrating
along Earth’s magnetic field lines.
→ Magnetosome synthesis begins with
insertion of magnetosome specific
proteins into the cytoplasmic membrane.
• Followed by invagination of the
membrane to form a vesicle
• Vesicle is filled with iron in the
oxidation state Fe (II)
→ Biomineralization proceeds through the
activities of the magnetosome proteins
• Includes an iron oxidase enzyme that
catalyzes formation of the Fe (III)
needed to form the magnetic minerals
→ Morphology of magnetosomes varies and
is species specific.
• Several are possible
• Square, rectangular, or spike-shaped
magnetosomes are most common.
ENDOSPORES
→ Bacteria produce specialized spores
called endospores.
→ Endospores: highly differentiated
dormant cells that function as survival
structures and can tolerate harsh
environmental conditions (extreme heat,
 radiation, chemical exposure, drying,
nutrient depletion)
• Prefix endo means “within”
→ Endospores are not reproductive
structures.
• They are the dormant stage of a
bacterial life cycle.
• Vegetative cell → endospores →
vegetative cell
→ Many microbes can form spores or spore-
like structures
• Myxospores of myxobacteria
• Arthrospores of actinomycetes
• Various forms of cysts
→ Endospores are exceptional for their
extreme resistance to environmental
challenges
• Only type of spore that can tolerate
high heat
• Is able to survive hours in boiling
water
→ Endospores can remain dormant for
hundreds and even thousands of years
only to germinate and grow when
conditions become favorable
→ Endospores are easily dispersed by wind,
water, or animal gut
• Endospore-forming bacteria are
widely distributed in nature
→ Endospores are only produced by two
groups of bacteria (both are gram positive
bacteria of phylum Frimicutes):
• Bacillales
• Clostridiales
→ These bacteria share an ancestor
• It is likely that the ability to form
endospores evolved only once
→ Best student endospore formers are in
the genus:
• Bacillus
• Clostridium
→ These bacteria are found widely in soil
and other environments
• Some are well known pathogens of
humans and other animals.
→ Endospore forming bacteria are a major
cause of food spoilage and foodborne
disease
→ Botulism, tetanus, and several foodborne
bacterial infections are caused by species
of endospore forming bacteria.
ENDOSPORE FORMATION AND
GERMINATION
→ Sporulation: process of cellular
differentiation that results in endospore
formation
→ During sporulation/endospore formation:
• A vegetative cell is converted through
a process of cellular differentiation
into a nongrowing, heat resistant, and
light-refractive structure.
→ Sporulation is triggered when some
nutrient becomes limiting.
• When the cell senses this, the
developmental process that converts
vegetative cell into an endospore
begins
• It is controlled by a complex
regulatory system.
→ An endospore can remain dormant for
years
→ When conditions are favorable growth, it
can convert back to a vegetative cell
rapidly through the process of
germination
→ Germination is triggered by the
availability of nutrients such as certain
amino acids or sugars
→ Germination occurs in three steps:
• Activation
• Germination
• Outgrowth
→ Overall process occurs in just a few
minutes.
• It is characterized by hydration of the
spore
• Results in loss of its heat and
chemical resistance, loss of specific
spore structures and the regeneration
of vegetative cell structures
• Ultimately the onset of vegetative
growth by binary fission.
ENDOSPORE STRUCTURE AND FEATURES
→ Endospores are visible by light
microscopy as strongly refractile
structures
→ Endospores are impermeable to most
dyes
• Occasionally they are seen as
unstained regions within cells that
have been stained with basic dyes
such as methylene blue.
→ To stain endospores:
• Special stains and procedures must
be used
• The classical endospore-staining
protocol uses malachite green and is
infused into the spore with steam.
→ Structure of the endospore differs
distinctly from the vegetative cell.
→ Endospore contains many layers absent
from the vegetative cell
→ Innermost region of the endospore is
called the core
• Contains DNA and ribosomes
• Develops from the cytoplasm of the
vegetative cell
→ Surrounding the core is the:
• Inner membrane: develops from the
cytoplasmic membrane of vegetative
cell
• Cortex: composed of peptidoglycan
• Outer membrane: special membrane
formed during sporulation (should not
be confused with the LPS).
→ Beyond the outer membrane is the
endospore coat:
• Composed of layers of spore-specific
proteins
→ Some but not all endospores also have • SASPs function as a carbon and
an outer proteinaceous layer called the energy source for the outgrowth of
exosporium. a new vegetative cell during
germination.
→ Endospore toughness and the reason
that endospores are highly refractile lies
in the dehydration of the core.
• Endospore core contains less than
one-quarter of the water found in the
vegetative cell

→ Dehydration increases heat and chemical

resistance.
• Causes enzymes in the core to
become inactive.

→ Dehydration of the core is facilitated by

the accumulation of a substance called
dipicolinic acid

→ Endospores also contain large amounts

of calcium (Ca2+), most of which is
complexed with dipicolinic acid.
→ The calcium-dipicolinic acid complex
forms about 10% of the dry weight of the
endospore and functions to bind water,
helping to dehydrate the developing
endospore.
• The complex also inserts between
bases in DNA which helps stabilize
DNA against heat denaturation.
→ Endospore core also contains high levels
of small acid-soluble spore proteins
(SASPs)
→ SASPs are only made during sporulation
process and have two functions:
• SASPs binds tightly to DNA in the
core and protect it from potential
damage from ultraviolet radiation,
desiccation and dry heat
o Ultraviolet resistance is conferred
when SASPs alter the physical
structure of DNA causing it to
become more compact
o Change in DNA causes it to be
more resistant to mutations and
damage.
SPORULATION CYCLE
→ Sporulation is a form of cellular
differentiation
• Many genetically directed events
occur during the conversion from
vegetative growth to sporulation.
→ Bacillus subtilis has been studied in
detail.
→ Conversion of a vegetative cell into an
endospore takes about 8 hours.
→ Begins with asymmetric cell division and
the formation of a forespore.
→ Engulfment of the forespore by the
mother cell results in the formation of the
outer membrane that surrounds the
developing endospore.
• The outer membrane forms from part
of the cytoplasmic membrane.
→ Key events in endospore formation such
as asymmetric cell division, cortex
 formation, and SASP production takes
place in a defined sequence and at
specific time.
→ Genetic studies of mutant of Bacillus
subtilis, each blocked at one of the stages
of sporulation.
• This indicates that more than 200
sporulation specific genes exist
• These genes are turned on and off in
a genetic program that governs
cellular differentiation.
CELL LOCOMOTION
→ Many microbial cells can move under
their own power.
→ Motility allows cells to reach different
parts of their environment.
• In nature, a new location may offer a
cell additional resources or protection.
FLAGELLA, ARCHAELLA, AND SWIMMING
MOTILITY
→ Many Bacteria are motile by swimming
due to a structure called the flagellum
(flagella)
• Analogous structure called
archaellum is present in many
Archaea.
→ Flagella and archaella are tiny rotating
machines that function to push or pull a
cell through liquid.
FLAGELLA AND FLAGELLATION
→ Bacterial flagella are long, thin
appendages (15-20 nm wide) free at one
end and anchored to the cell at the other.
→ Flagella can be stained and observed by
light microscopy or electron microscopy.
→ Flagella can be anchored to a cell in
different locations:
• Polar flagellation: flagella are
attached at one or both ends of a cell
o Lophotrichous: group of many
flagella (tuft) can arise at one end
of a cell, a type of polar
flagellation.
o Amphitrichous: tuft of flagella
emerges from both poles of the
cell.
• Peritrichous flagellation: flagella are
inserted around the cell surface
→ Flagella do not rotate at a constant speed
• They increase of decrease their
rotational speed in relation to the
strength of the proton motive force
→ Flagella can rotate at up to 1000
revolution per second
• Supports a swimming speed of up to
60 cell lengths/sec
 • Bacterium is actually moving over
twice as fast relative to its size as the
faster animal.
→ Swimming motions of polarly and
lophotrichously flagellated organisms
differ from the peritrichously flagellated
organisms
→ Peritrichously flagellated organisms
typically move slowly in a straight line,
stop, then new direction.
→ Polarly flagellated organisms move more
rapidly and continuously, and some are
able to reverse their direction.
FLAGELLA STRUCTURE AND MOTILITY
→ Bacterial flagella are rigid and helical.
→ Filament: main part of the flagellum
• Composed of many copies of a
protein called flagellin.
• Amino acid sequence of flagellin is
highly conserved in Bacteria
• Suggests that flagellar motility evolved
early and has deep roots within this
domain.
→ Hook: wider region at the base of the
filament that consists of a single type of
protein and connects the filament to the
flagellum motor in the basal body.
→ Flagellum motor: reversible rotating
machine composed of more than 25
proteins and anchored in the cytoplasmic
membrane and cell wall.
→ Motor consists of a central rod that
passes through a series of rings.
→ In gram negative bacteria:
• L ring: outer ring anchored in the
outer membrane
• P ring: second ring anchored in the
peptidoglycan layer.
• MS and C rings: third set of rings
located within the cytoplasmic
membrane.
→ In gram positive bacteria which lacks an
outer membrane only the inner pair of
rings is present (P, MS, and C ring)
→ Stator: surrounds the inner rings and
anchored in the cytoplasmic membrane
and the peptidoglycan layer.
• Composed of Mot proteins.
→ Export apparatus: a type III secretion
system that facilitates synthesis of the
flagellum.
• On the cytoplasmic side of the MS
ring.
→ Flagellum motor contains two main
components:
• Rotor: consists of the central rod and
the L, P, C, and MS rings.
• Stator: compromised of Mot proteins
which surround the rotor and function
to generate torque.
• Collectively these structures make up
the basal body.
→ Rotation of the flagellum occurs at the
expense of the proton motive force.
• It is thought that rotation is caused by
a type of “proton turbine” process.
→ In this model, proton translocation
through channels within the stator
complex cause the MS ring to rotate,
driving rotation of the attached rod and
flagellum.
• Protons flowing through the Mot
proteins exert electrostatic forces on
helically arranged charged residues
on rotor proteins and cause MS rings
to rotate.
→ The L and P rings act like bushings within
which the rod rotates.
→ About 120 protons are translocated by
each rotation of the flagellum.
→ Rotational speed is set by the proton flow
rate through the Mot proteins.
• Is a function of the intensity of the
proton motive force.
→ The flagellar motors of different microbes
are able to generate different amounts of
torque.
• Causes significant differences in
swimming speed
• Such adaptations are driven by
adding or subtracting subunits from
the stator and C ring
FLAGELLAR SYNTHESIS
→ Several genes encode the motility
apparatus of Bacteria
• In Escherichia and Salmonella
species, over 50 genes are linked to
motility.
→ These genes encode the structural
proteins of the flagellum and motor
apparatus.
• Also encode proteins that export the
structural proteins through the
cytoplasmic membrane to the outside
of the cell and proteins that regulate
the synthesis of new flagella.
→ A flagger filament grows from its tip.
→ MS ring is synthesized first and is
inserted into the cytoplasmic membrane.
→ Then the anchoring proteins are
synthesized along with the hook and cap
before the filament forms.
→ Flagellin molecules are synthesized in the
cytoplasm.
• Is exported through the export
apparatus.
• Export apparatus shuttles flagellin
molecules into a 3 nm channel that
runs through the basal body and
hollow filament.

→ The flagellin subunits are ultimately

passed to the end of the filament where
they attach to the terminus of the growing
flagellum

→ Cap proteins helps flagellin molecules

assemble.

→ Approx. 20,000 flagellin molecules are

needed to make one filament

→ Flagellum grows more or less

continuously till reaches final length

→ Broken flagella still rotate and can be

repaired with new flagellin units.

ARCHAELLA
→ In Archaea, swimming motility is also
widespread.
• Is driven by the rotation of their
flagellum analog the, archaellum.
→ Archaella proteins are unrelated to those
of flagella.
• Evolutionary terms are most closely
related to the type IV pili.
→ Archaella are somewhat smaller than
flagella (10-13 nm in width)
• They are slightly larger than type IV
pili.
→ Archaella are not hollow and are
assembled from their bases.
• Flagella are hollow and assembled
from their tips.
→ Archaellar rotation is driven by ATP
hydrolysis.
• Flagellar rotation is from proton
motive force,

→ Archaellar motor is structurally simpler

than the flagellar motor.

→ Only 7-12 genes are required to encode

major proteins that make up the
archaellar motor
• More than 25 genes for flagellar
motor.

→ Swimming Archaea, such as

Halobacterium, show that they swim at
speeds only about one-tenth that of
Escherichia coli.

→ In general, Archaea swim much more

slowly than Bacteria.
• Could be due to the smaller diameter
of the archaellum.
• Also due to difference in the
constructure of motor and how it is
powered

→ Some Archaea have found a way to

overcome these limitations.
• Example: Methanocaldococcus
swim nearly 50 times faster cells of
Halobacterium and 10 times faster
than cells of Escherichia coli.

SURFACE MOTILITY