MICB 201 Chapter 2: Learning Objectives/Outcomes (2010W T1)
Define the terms: coccus, bacillus, spirillum, vibrio, hypha, mycelium and plasma in the context of
prokaryotic cell shape.
Coccus – berry/sphere
Bacillus – rod
Spirillum – spiral/helix/coil
Vibrio – comma shaped (rod with bend)
Hypha – web/thread
Mycelium – many hypha, means fungus
Plasma – fluid – indicates a very variable/irregular shape
Compare Bacteria and Archaea with respect to cell shape, multicellularity and cellular differentiation.
You basically can’t tell a bacterium from an archeael organism without genome analysis. Basically, you
can’t tell them by cell shape most of the time. Only bacteria can differentiate, Archaea cannot. They
can both be multicellular if they want.
Explain the importance of cell SA:V relationships in nutrient acquisition and waste disposal.
SA:V is important. Surface area is related to how fast nutrients are acquired and how fast waste is
disposed of. Volume is related to how much nutrients are required and how much waste is produced.
As the size of a cell increases, the SA:V decreases because volume increases more quickly than surface
area.
Estimate the SA:V ratio of a cell given relevant information.
Describe how some prokaryotes modify their effective surface area to volume ratio using membrane
invagination and vacuole formation.
Some cells are very large. That means their SA:V is small relative to other smaller cells. Therefore they
must seek to increase it. They can do this by using vacuole formation, which decreases the effective
volume of the cell because the vacuole may be metabolically inactive. Invagination increases surface
area that the cell can use. Stalks may also be used.
Describe the composition and organization of the prokaryotic cytoplasmic membrane.
The bacterial and archaeal cytoplasmic membrane is a bilayered structure about 10 nm thick composed
of phospholipids just like eukaryotes. Each half is called a leaflet. Embedded in the lipid bilayer are
proteins that are integral and may span both leaflets or just one. 50:50 lipid to protein by weight but
30:1 phospholipids molecules to a protein. The phospholipid has a head that may be an amino acid or
sugar bonded to glycerol-phosphate. The glycerol is actually connected to phosphate and also to two
hydrocarbon tails (fatty acids). All Archaea use an ether group –C – O – C- to connect the glycerol to the
fatty acid tails. Most bacteria use an ester linkage (-COO-) to do this, but some may use the ether. The
hydrocarbon tails of bacterial membrane phospholipids are unbranched most of the time. In Archaea
there are methyl branches off the main hydrocarbon chain.
Appreciate that the molecular structures of bacterial and archaeal phospholipid exhibit differences.
As said earlier, all Archaea use an ether linkage which is –O – C – O- to link glycerol phosphate to the
fatty acid hydrocarbon tails while most Bacteria/Euk use an ester linkage instead. Archaea also have
branching (methyl groups) and Bacteria/Euk usually do not have branches.
Also, the stereochemistry of the glycerol phosphate group. Bacteria and euk phospholipids use the D
stereoisomer of the glycerol while Archaea use the L stereoisomer.
Describe the mechanisms of passive and active transport of nutrients in prokaryotes and the different
ways energy can be supplied to the later process (eg. ABC transporters, H+ gradients)
Permeases are the proteins that are used in cytoplasmic membrane. Diffusion occurs which is passive
transport. This is for small molecules that tend to be nonpolar, uncharged. Oxygen, carbon dioxide and
water pass through the membrane directly. This is because phospholipids are not chemically bonded to
each other e.g. covalently. Large or charged chemicals must also pass through into or out of the cell.
Permeases are the proteins used for this purpose. Each permease transports a different chemical. That
is, permeases are specific. Permeases may be for facilitated diffusion/passive transport or for active
transport. For facilitated diffusion, the molecule binds to the permease and it changes conformation.
Then it goes through. Molecules can just as easily leave the cell as entering using the permease.
However, backward transport is generally prevented by consuming the molecule that needs to be
transported upon entry.
Prokaryotes usually live in hypertonic solutions. Chemicals/nutrients must be accumulated inside the
cell. In order to work against the concentration gradient, the cell uses active transport methods. Active
transport is the utilization of energy to move a chemical from an area of low to high concentration. As
with facilitated diffusion, a chemical-specific transmembrane permease must be used.
One method of active transport in prokaryotes is via ATP consumption. This involves ABC transporters
which are ATP-Binding Cassette Transporters. These are found in all 3 domains. They are important.
Consists of a transporter protein that is transmembrane and forms a pore through the CM. On the
cytoplasmic end there is a nucleotide-binding site where ATP can bind to be used. On the other side, it
depends. In Gram negative cells there is a OM so you can have periplasmic proteins floating around.
For Gram positive cells the proteins are attached to the membrane, anchored to the membrane. These
binding proteins bind to the chemical and pass them on to the transmembrane transporter protein in
the CM. Conformational changes in the transporter protein promoted by energy from ATP consumption
move the chemical into the cell.
Another method is cotransport. This is also active transport and uses a concentration gradient rather
than consuming ATP. Protons can be transported from the periplasm across the CM to the cytoplasm.
There are no periplasmic binding proteins in this process.
Aquaporins are proteins that move water across bilayers faster than simple diffusion allows. They are
common in many eukaryotes including plants and mammals. They are not that common in microbes.
Describe the nature of the prokaryotic cytoplasm and why the shape of phospholipids is important in
delineating this compartment.
The cytoplasm has loads of ribosome and contains the nucleoid. There are usually no membrane bound
compartments. The shape of phospholipids determine the shape of the cytoplasm. The cytoplasm is
both a concentrated solution of protein and a dense suspension of ribosomes, mRNA, tRNA and other
random molecules. 10s of thousands ribosomes per cell. Proteins account for 50% of the cell dry
weight. Phospholipids have two HC tails making them like a cylinder. In water, these phospholipids
pack into vesicles containing an internal aqueous compartment. If phospholipids had one tail then bad.
Obviously. Allows for internal aqueous compartment.
Explain what the terms Gram-positive and Gram-negative mean.
They refer to how bacterial cells show up when fixed to a slide and stained with two dyes. It is related
to the cell wall structure but not much can be implied. It is basically a method of categorizing bacterial
cells based on how they show up under the dyes. Gram positive cells stain purple while Gram negative
cells stain red.
Without resort to detailed molecular structures, describe the composition, structure and function of
peptidoglycan in Gram-negative and Gram-positive bacteria.
Peptidoglycan is a polysaccharide that exists in the cell wall of both Gram positive and Gram negative
cells. It is not found in any domain other than Bacteria. PG is a huge mesh like molecule composed of
two parts – the glycan chain and the tetrapeptides that connect the glycan chains. The glycan chain is
made up of two subunits that are joined by an O-glycosidic bond. They are called NAM and NAG which
stand for N-acetylmuramic acid and N-acetylglucosamine. The O-linkages are strong covalent bonds.
The tetrapeptide is a sequence of 4 AA that are joined by strong covalent bonds in peptide linkages. The
N-acetylmuramic acids of the glycan chains are crosslinked to each other through amino acid 3 and
amino acid 4 of their tetrapeptides. Amino acid 4 is always alanine and Amino acid 3 is important
because it must possess a free NH3+ group – the crosslink is formed from the –COO- group of Ala (AA4)
and the –NH3+ group of AA3. In Gram negative bacteria, the tetrapeptides are directly cross linked to
each other. In Gram positive bacteria, there may be up to 5 additional amino acids in a peptide
interbridge. Gram positive PG has a higher degree of cross linking than Gram negative PG. As much as
80% of the NAMs in G+ bacteria can be crosslinked while less than 50% of the NAMs are cross linked in
G- bacteria and can be as low as 20%. 20% to 50% cross linkage in the NAMs for Gram negative bacteria,
up to 80% for Gram positive bacteria and Gram positive bacteria have an interbridge b/t NAMs that is up
to 5 AA long. The AA of the tetrapeptide alternate L and D configurations. Perhaps to prevent
degradation.
Without resort to detailed molecular structures explain the differences between the composition and
structure of Gram-negative and Gram-positive cell walls.
Basically, the Gram-negative bacteria has an outer membrane (which contains porins and possibly
permeases), then underneath some space, then the peptidoglycan layer which is like 3-5 layers thick and
then some more space btw this is in the periplasm and then the cytoplasmic membrane.
In Gram positive bacteria, there is no OM but there is peptidoglycan layer which is very thick. CM as
usual. About 30% of cultured bacteria are Gram positive. Gram positive cell wall layers can have as
many as 20 layers of PG producing a structure hundreds of nm thick. Gram positive cell walls contain
polymers of phosphorylated sugar alcohols based on ribitol and glycerol and these are called teichoic
acids. They are further classified into just teichoic acids or lipoteichoic acids. If the teichoic acid
polymer is anchored to a sugar in the headgroup in a cell membrane phospho or glycolipid, it is called a
lipoteichoic acid. If it is linked to the peptidoglycan NAM, then it is simply called teichoic acid. Both
poke through the PG mesh and into the environment, not sure what they are for though.
Unlike previously believed, Gram positive cells do have a periplasm between the PG and the cell
membrane.
The Gram negative cell wall is the most common bacterial cell wall type, accounting for about 50% of
cultured bacteria. Gram positive cell walls only have about 1 to 2 layers of peptidoglycan. External to
the PG is the outer membrane which is a bilayer structure that is made out of lipopolysaccharide,
phospholipid and protein. The outer membrane contains phospholipids but only on the inner leaflet. In
the outer leaflet there are other lipid molecules called LPS – lipopolysaccharide. There are three regions
that the LPS is divided into. Lipid A, R-core and O-polysaccharide. Lipid A is made of several
hydrocarbon tails and is the thing that is inserted into the membrane. The R core is a negatively charged
that is made of many sugars. The O-polysaccharide is what extends outwards from the cell and attached
to the R-group. Obviously made of sugars, but uncharged unlike the R-core which is made out of
charged sugars (negative). Specific sugars can differ and sometimes the O-polysaccharide is not there.
The outer membrane contains proteins – lipoproteins and porins. Lipoproteins are proteins with
hydrocarbon tails (the lipid part). They HC tail is embedded in the outer membrane and the protein part
is covalently bonded to the peptidoglycan. Thus the lipoprotein anchors the outer membrane to the
peptidoglycan. The outer membrane lipid bilayer is even less fluid and less permeable than the CM.
This is due to stronger lateral interactions b/t the lipopolysacchardide molecules than between
phospholipids. There are also Calcium 2+ and Magnesium 2+ ions present in the outer leaflet of the
outer membrane which help stabilize the negative charge of the LPS R-core. Therefore there are ionic
bonds which stabilize the membrane and make it less fluid. It links the adjacent LPS molecules. Second,
LPS has more than 2 HC per tail (sometimes 6) resulting in increased hydrophobic effect and increased
lateral interactions (IDID) because of the increased SA.
State the prevalence and distribution of the Gram-positive and Gram-negative cell wall types in the
bacterial domain.
30% of cultured organisms are G+, 50% G-.
Without resort to detailed molecular structures, describe the composition, structure and function of the
Gram-negative outer membrane.
The Gram negative outer membrane is made out of phospholipids and LPS which stands for
lipopolysaccharide molecules which are made out of many hydrocarbon tails (more than 2), R-core
(negative charge) and O-polysaccharide. Same width as CM which is about 10 nm. Less fluid because of
the lateral interactions (more hydrocarbon) and also more ionic because MG2+ and Ca2+. There are
porins and lipoproteins in the Gram negative outer membrane. The lipoproteins have two hydrocarbon
tails which are embedded in the inner leaflet of the outer membrane and the protein is covalently
bonded to PG which means the function is to attach the OM to the PG.
Other than lipoproteins, there are porins that are transmembrane proteins. Because the OM is such an
effective barrier against permeation, need some porins for transport. Porins are for small nutrient
molecules (charged and uncharged) and this is passive/facilitated transport – facilitated diffusion.
Porins form water-filled channels or pores in the OM. Some porins are general and allow passage of all
chemicals that are small enough. Others are chemically selective, specific and therefore only allow
certain types of chemicals. This selectivity is determined by the AA lining the interior of the porin
molecule in the OM. Different G- organisms possess porins with different properties.
The function of the G- OM is protective in nature. Provides additional permeability barrier to protect PG
and CM from harmful chemicals both small and large in the environment. Lysozyme is a good example
of a large molecule. Because PG is exposed to environment, G+ bacteria are sensitive. So G- have an
advantage. The lysozyme does not readily penetrate b/t the gaps that open between the lipids in the
OM nor does it generally pass through the porin channels in the OM.
Describe the nature of the periplasm in Gram-negative and Gram-positive bacteria.
In both G+ and G- bacteria, there is something called a periplasm which is a water filled compartment.
The periplasm is a dense aqueous solution of proteins. The protein concentration is so high that the
periplasm is almost gel like. Several proteins make up this gel, serving multiple functions.
Describe the essential features of periplasmic and extracellular protein secretion in Gram-positive and
Gram-negative bacteria.
Proteins cannot simply diffuse across the CM and into the periplasm because they are large charged
molecules. They must be transported using transport proteins. This is active transport and involves the
consumption of ATP. Bacteria also transport proteins out of the cell (extracellularly) to serve various
functions.
In Gram positive bacteria, the bacteria simply have the protein actively transport from the cytoplasm to
the periplasm across the CM and then it simply diffuses through the PG and out. Not clear why or how
periplasmic proteins are retained in the periplasm for them to function there.
In Gram negative bacteria, same thing but then it also needs to go across the OM. There are various
models that have been proposed to show how Gram negative cells transport proteins extracellularly.
One model says that they are first transported to the periplasm and then into the environment via
another protein complex in the outer membrane bilayer. Another model says that instead of going into
the periplasm it just goes straight out of the cell by having a protein complex that spans the periplasm,
CM and OM at once. Periplasmic proteins stay in the periplasm because there is the OM.
In these methods of transport to out of the cell or to the periplasm, the protein obviously needs to be
unfolded during/before transport. Active transport.
Describe the unique features of Mycoplasma cell structure.
Most Bacteria have a cell wall but some do not, such as the Mycoplasmas. They are related to Gram
positive bacteria and they are parasites to both plants and animals. They lost their cell wall through
reductive evolution. The loss of these energy expensive structures is an adaptation to their parasitic
lifestyle and has enhanced their reproductive rate in the protected, osmotically compatible host
environment since they don’t need to make the peptidoglycan. They are not as osmotically stable
obviously without a PG as bacteria w/ cell walls, they are more osmotically stable than protoplasts. So
that means their CM is stronger, although not really sure how. Also, mycoplasmas are the smallest of
bacteria with cell diameters around 200 nm. Less force on the CM.
Distinguish between osmotic lysis and plasmolysis and the conditions under which each occur.
In hypotonic solutions, [solute] in the cell > [solute] in the solution. Therefore, osmosis => water rushes
in and cell will lyse (osmotic lysis) if there is no protective barrier such as a cell wall (e.g. PG). Lysozyme
catalyzes the hydrolytic cleavage of the O-glycosidic covalent bonds that link NAM and NAG. So if
remove PG first with lysozyme, osmotic lysis can occur. PG doesn’t protect cell from plasmolysis. If the
PG is disrupted (with lysozyme or something) under isotonic conditions cell lysis does not occur but
rather a protoplast is left behind – a membrane bound structure that is osmotically fragile. It is usually a
sphere since that is the ‘default’ shape in nature. Therefore the PG helps retain the shape (rod, etc).
Describe the general features of the cell walls of the Deinococcus bacterial group, the green nonsulfur
bacterial group and the Planctomyces group.
Deinococcus – have OM like G- but OM lacks LPS. Some stain G+ some G-. Not really understood, may
have to do with S layer. Deinococcus may or may not have S-layer, OM is non-LPS based, some PG
layers, CM.
Green nonsulfur – Green nonsulfur bacteria (and relatives) have a Gm+ cell wall structure, sometimes
with S layer, but do not usually stain positive. S-layer may be there, some PG may be there, CM.
Planctomyces – these Bacteria don’t have PG. Instead, have many copies of the same protein
molecule joined by covalent bonds as a rigid layer. This protein layer seems to constitute the entire cell
wall.
Describe the composition and structure of bacterial capsules, S-layers, pili and flagella.
Surface (S) layers are composed of many copies of the same protein arranged in a geometric pattern
anchored to an underlying cell wall component. Tetragonal or hexagonal. In Gm+ bacteria, S-layers are
anchored to the PG while in Gm- O-polysacc and teichoic acids poke through the holes in the S-layer and
are exposed on the cell surface. S layers do not provide mechanical strength as they are not joined
together covalently. The function is unknown.
If a cell has a capsule, it is the outermost layer covering a S-layer if present. Chemically, capsules fall
into 2 types. Polysaccharide capsules and polypeptide capsules. Polysacc capsules are chains of a single
type of sugar. Polypeptide capsules are chains of a single type of AA. In Gram negative bacteria,
capsular polysaccharide is frequently anchored to the cell by replacing the O-polysaccharide of the LPS
or via phospholipid. In Gram positive bacteria, capsules are linked to the NAM and NAG sugars making
up PG. Function of capsules? To protect bacteria from dessication/drying – capsular material is
hydrophilic and binds lots of water via H-bonding. 95% weight of isolated capsular material are water.
Capsules can protect bacteria from phagocytosis – “slippery.” Capsules can be involved in attachment
to surfaces.
Pili are filamentous projections from the cell surface. There can be many or not depending. They can be
very long. Gram negative pili are made out of major proteins called pilins. The subunits form chains
wrapped around each other like the structure of ropes. Like actin filaments. But pilins for Gramnegative
pilus.
Pili of Gram positive bacteria have pilin proteins that are covalently bonded and the entire structure is
covalently bonded to un crosslinked peptide of the PG – these pili are not retractable.
Retractable pili in Gram negative bacteria. There is a basal body embedded in the CM and have the pili
(noncovalently bonded) wrapped around each other like a double helix, going out of the peptidoglycan
and the OM.
Non retractable pili in Gram negative bacteria. Attached to the protein in the OM – called OM protein
‘anchor.’
Retractable pili in Gram positive bacteria. Remember, covalently bonded unlike in Gram negative
bacteria. Bonded to the basal body in the CM. In nonretractable pili in Gram positive bacteria, again
covalently bonded but attached to the peptidoglycan. It is covalently bonded to un cross-linked peptide.
There is no double helix in the nonretractable pilus of Gram positive bacteria, just covalently bonded
subunits pilins.
Summary – covalent bonds do not exist in all retractable pili and in all Gram negative bacteria pili. Only
in the Gram positive nonretractable pilus that there are covalent bonds.
Flagella – one way that bacteria move is by swimming, using flagella. These are long filamentous
appendages. Monotrichous bacteria have only one flagellum. Peritrichous bacteria have many. Flagella
are made out of several different proteins. Made out of flagellin subunit proteins. The filament is
attached via a hook protein to a rod protein which is embedded in the membrane and wall by several
protein rings. Transport of H+ drives the flagellum rotation. In the CM there’s a rod protein embedded.
It spans the peptidoglycan layer and the OM as well. The rings surround the rod protein M, S, P protein
ring. In the environment the rod protein is attached to a hook which is made out of hook proteins which
is attached to a filament made out of flagellin. Some diff. b/t G+ and – but mainly number of rings.
There is also a basal structure embedded in the CM that surrounds some rings and obviously the rod
protein in the CM. There is the MS and C ring in the CM. There are Fli proteins which are the motor
switch and Mot proteins through which protons are transported from the periplasm to the cytoplasm.
Describe the general features of endoflagellar swimming in spirochetes and gliding locomotion including
gliding in Mycoplasmas.
Endoflagella – there are endoflagella in spirochetes. There are two in the periplasm between the PG
and outer membrane (this spirochetes is Gram negative). Endoflagella from the two poles overlap for
much of the cell length. Spirochetes are spiral Bacteria with a Gram negative cell wall. All members of
the Spirochete group has endoflagella and are confined w/n the periplasm between the PG and the OM.
They are anchored in the CM at the poles of the cell and spiral outside the PG but inside the OM,
overlapping at the midpoint. The cell body and endoflagella thus form a double helix, winding around
each other. The endoflagella rotate inside the periplasm. This somehow allows the entire spiral cell to
rotate and corkscrew through the water.
Gliding motility – bacteria move along a surface. Observed in Purple Bacteria group, Cyanobacteria,
Green sulphur and some Green nonsulfur. There are at least two mechanisms – pilus based and slime
based. Slime based – excrete slime onto surface, slime hydrates and increases in volume, therefore
pushing the bacteria along. Pilus based – pilus extend (many) attach to surface. Retract – this pulls the
bacteria along.
State the prevalence and distribution of the different forms of locomotion in the bacterial domain.
Describe the general structural features of the different cell wall types elaborated by the Archaea.
Compare the properties of bacterial and archaeal S-layers in terms of structure and function.
Compare the properties of bacterial and archaeal flagella and pili in terms of structure and action.
Most Archaea are unicellular. Only a few species are filamentous. Many Archaea lack a rigid cell wall
polysaccharide so they can have irregular/pleomorphic shapes.
There is tremendous diversity in archaeal cell wall structure. In Bacteria, S-layers if present are not part
of the cell wall. In Archaea, S-layers are an integral cell wall component. In some organisms S-layers are
the cell wall. There are stronger interactions between the S-layer protein subunits of archaeal S-layers
and between the S-layer proteins and underlying cell components.
Also, bacteria use PG while Archaea use another rigid polysaccharide component like
pseudopeptidoglycan.
Archaea can also make various cell surface appendages including flagella and pili. They can use flagella
to swim. However, the archaeal flagellum does not rotate and the basal structure lacks protein rings.
In Bacteria, use transport of H+ to power flagellum. In Archaea, ATP is consumed instead.
Describe the general features of endoflagellar swimming in spirochetes and gliding locomotion including
gliding in Mycoplasmas.
Mycoplasmas which are cell wall-less parasites, have gliding motility. Motile cells are often shaped like
bowling pins and move in the direction of the head. There are two mechanisms. Fast gliders walk on
tiny protein legs localized to the next region with movement of the legs in a centipede-like motion. ATP
is consumed in this process providing the energy for motility. The head of slow gliding Mycoplasmas is a
narrow constriction at the front of the cell, a terminal organelle. There are proteins and they attach to
the surface. Extension and contraction of the terminal organelle and simultaneous release of the
proteins from the underlying surface during the cycle yields an inchworm-like movement.
Appreciate bacteria as organisms that sense and respond to their environment by describing the classic
experiments establishing the nature of flagellar-based chemotaxis.
Chemotaxis – behavioural response to sensing a chemical concentration difference in the environment.
Many motile prokaryotes exhibit chemotaxis. Can move toward attractants (positive chemotaxis) or
away from repellents. Nutrients are attractants, waste products are repellents.
In the ABSENCE of a concentration gradient, motility does not result in net movement in any particular
direction. In the PRESENCE of a nutrient concentration gradient, net movement occurs in a specific
direction – chemotaxis.
Chemotaxis in unicellular peritrichously flagellated bacteria like E. coli. In these motile bacteria, rotation
of all the flagella is in one direction (e.g. CCW) causing them to come together in a bundle and act in a
coordinated manner. The bacterium is thus propelled forward – this is called a run. Rotation of the
flagella in the opposite direction (CW) causes the flagellar bundle to FLY APART – this is a tumble.
Random change  direction is changed randomly and bacterial flagella go CCW again for another run.
Bacteria sense concentration temporally (over time) rather than spatially (at both poles). Bacteria can
tumble faster if senses its going in the wrong direction. Bacteria can inhibit tumbling (longer runs) if
going in the right direction.
Describe the difference in the motile behaviour of a peritrichously flagellated unicellular bacterium in
the presence and absence of a nutrient concentration gradient and explain the reason for this difference
in behaviour.
Monotrichously flagellated cells do not tumble. Rather, rotation of the flagellum in one direction pushes
the cell in a smooth running motion while rotation in the other pulls the cell backward in an erratic way
that causes it to change direction randomly.
Another way that monotrichous flagellated cells move is by only turning their flagella in one direction.
During a tumble, it stops and waits for Brownian motion to turn it.
• Solve problems similar to those in the Chapter 2 Questions and Problems handout (VISTA).