MBIO 1010 / Topic 2b: Cell Wall

What does the bacterial cell wall look like? Is it true that all bacterial cells have a cell wall?

Rigid layer. Not all bacterial cells have a cell wall.

What are the four roles of a cell wall?

• Protects cell against osmosis.

• Protects cell against toxins.

• Assists in pathogenicity.

• Helps shape the cell.

How permeable is the bacterial cell wall? What explains its level of permeability?

Not as permeable as the cell membrane. It’s full of holes, porous to most small molecules.

Just a review, where does water move from or into a cell in an isotonic, hypotonic, and hypertonic solutions? What happens in each situation thereafter?

• Isotonic? No net movement.

• Hypotonic? Water moves in, causing cell to swell.

• Hypertonic? Water moves out, causing cell to shrivel.

What does the cell wall do in regard to protection against osmosis?

The cell wall prevents osmotic lysis, wherein the cell bursts.

What does the cell wall do in regard to protection against toxins?

The cell wall protects cell against toxic substances, such as large hydrophobic molecules. While the cell wall doesn’t protect as much as the cell membrane, it still protects.

What are the two ways the cell wall does in regard to assistance in pathogenicity?

• Bacteria modify their cell walls to evade host immune system from identifying them.

• Cell wall helps bacteria stick to surfaces.

What does the cell wall do in regard to shaping the cell?

Once you remove the cell wall, the bacterium becomes coccus.

What are the two different types of bacteria based on the cell wall? Briefly define each.

• Gram positive, with their simple cell wall consisting of many layers of peptidoglycan.

• Gram negative, with their complex cell wall consisting of one layer of peptidoglycan and an outer membrane.

What does peptidoglycan look like, and what does it do? What is it made of?

Macromolecule acting as a rigid layer, providing strength to the cell wall. It is made of glycan tetrapeptide repeating units.

What is the glycan chain of peptidoglycan made of?

Two alternating sugars: N-acetylglucosamine (G or NAG) and N-acetylmuramic acid (M or NAM).

What is NAM attached to?

A tetrapeptide.

What holds the sugars together? What holds the amino acids together?

• Sugars held by ß-1,4 glycosidic bonds.

• Amino acids held by peptide bonds.

What is the glycan tetrapeptide unit composed of?

NAG, NAM, and the tetrapeptide.

Are all peptidoglycan structures exactly the same in all bacteria? Briefly explain why or why not.

No. Different gram positives and gram negatives have different peptidoglycans.

What is common in all structures of peptidoglycan, and what differentiates them from each other?

Common: All have NAG and NAM, with the latter holding a tetrapeptide that can cross-link an amino acid at position 3 to an amino acid at position 4 of another tetrapeptide.

Difference: All vary in tetrapeptide composition, cross-links, and or interbridge.

What are the two features that make peptidoglycan (and thus the cell wall) difficult to break down?

• Its ß-1,4 glycosidic bonds are less usual and stronger than α bonds in some other glycans, like starch and glycogen.

• Usual proteins are made of L-amino acids, cleaved off by most peptidases. Peptidases can’t easily cleave off the tetrapeptide because of the presence of unusual D-amino acids.

What is the tetrapeptide order in E. coli’s peptidoglycan?

L-alanine, D-glutamate, diaminopimelic acid (DAP), and D-alanine.

In E. coli, what position on NAM is DAP? How about D-alanine?

Position 3 and position 4, respectively.

What is the cross-link in E. coli’s peptidoglycan? How much cross-linking is there in E. coli’s peptidoglycan? What does the structure of peptidoglycan look like in the end?

DAP linked to D-alanine of another tetrapeptide on an adjacent peptidoglycan, with linking occurring in approximately 10–20% of tetrapeptides. This forms a net-like armour around the cell.

What is the tetrapeptide order in S. aureus’ peptidoglycan?

L-alanine, D-glutamine, L-lysine, and D-Alanine.

In S. aureus, what position on NAM is L-lysine? How about D-alanine?

Position 3 and position 4, respectively.

What is the cross-link in S. aureus’ peptidoglycan? How much cross-linking is there in S. aureus’ peptidoglycan?

L-lysine linked to a pentaglycine peptide interbridge, which brings L-lys to D-alanine of an adjacent peptidoglycan, with linking occurring in approximately greater than 90% of tetrapeptides.

What shape is peptidoglycan when viewed surrounding the cell? What does this allow?

Helical, allowing three-dimensional crosslinking.

Why is peptidoglycan important to us, humans?

The vast majority of bacteria have peptidoglycan, and it’s totally different from the cell wall of other cellular organisms. Heck, we humans don’t have peptidoglycan because we don’t have a cell wall. Since it is only found in bacteria, we can devise chemicals specifically damaging peptidoglycan without hurting us - a weapon against bacteria causing illnesses.

Provide two chemicals that we have made to combat against bacteria by taking advantage of their peptidoglycan.

Lysozyme and penicillin.

What is lysozyme? Where can you usually find it?

Enzyme cleaving ß-1,4 glycosidic bonds between NAM and NAG, damaging bacteria. It is commonly found in tears, saliva, and nasal secretions.

What is penicillin? What does it do?

Antibiotic preventing crosslink formation by inhibiting cross-linking, the mesh-like armour cannot be built. Growing bacterial cells burst due to osmotic pressure.

What do gram positives have as well in their cell wall, other than peptidoglycan?

Teichoic and lipoteichoic acids.

Where can you find teichoic acids, and what is its function?

Attached to peptidoglycan, extending outward, it helps give surface its negative charge.

What is a lipoteichoic acid, and what is its function?

Teichoic acid connected to a lipid at one end, linking peptidoglycan to cytoplasmic membrane.

What can you find “beyond” the gram negative cell wall?

An outer membrane.

How do we define the periplasmic space for gram negative cell walls?

The outer leaflet of the cytoplasmic membrane to the inner leaflet of the outer membrane.

What kind of phospholipids are found in the outer membrane of gram negatives? What differentiates the outer membrane (OM) of gram negatives from the cytoplasmic membrane (CM) of gram negatives?

Common: Ester phospholipids.

Different: OM’s leaflet mostly consists of lipopolysaccharide (LPS). OM also has different proteins from CM proteins.

What are the three parts of LPS?

Lipid A, core polysaccharide, and O-specific polysaccharide.

What is lipid A in LPS made of? Is it hydrophobic or hydrophilic? Which part of lipid A faces inward?

Sugars, phosphates, and fatty acids. It’s amphipathic, having the fatty acids face inward and the sugars facing outward.

What is core polysaccharide in LPS made of?

Sugars.

What is an O-specific polysaccharide made of? What’s another name for O-specific polysaccharide?

Also known as O-antigen, it is a long chain of sugars, varying in length and composition among gram negatives.

What is an antigen?

Substance detected by immune system, in order for immune system to defend.

What is LPS also called? Why is it called as such?

It is also called endotoxin, “endo” because it is made within the OM, and “toxin” because the lipid A portion is toxic to animals in an extreme amount.

When is LPS typically released? What common disease symptoms are caused by LPS release? What common gram negative pathogens release LPS?

When a Gram negative dies. Its OM breaks apart and LPS is released. Diarrhea, vomiting, and shock are common symptoms. E. coli and Salmonella are common pathogens.

What’s a porin? What is it made of? What does it do?

Protein trimer (made of three protein subunits), forming a channel through the outer membrane, allowing diffusion of small hydrophilic molecules.

What three proteins are commonly found in the periplasm?

Substrate-binding proteins, hydrolytic enzymes, and biosynthetic enzymes.

What do substrate-binding proteins do?

Bind substrates and move them to cytoplasmic membrane for transport into the cell, e.g. ABC transport.

What do hydrolytic enzymes do? Give an example of a hydrolytic enzyme and their specific function.

Degrade foreign substances, e.g. endonucleases destroying small foreign DNA.

What do biosynthetic enzymes do? Give an example of one and what it does.

Assist cell wall synthesis, e.g. penicillin binding proteins cross-linking peptidoglycan, accelerating cell wall synthesis.

Why is penicillin binding proteins called as such?

Penicillin molecules bind to these proteins, inhibiting their activity, ultimately leading to cell death.

In contrast to bacterial cell walls, what do archaeal cell walls not contain? What do they contain? Do they have an outer membrane?

Don’t: They don’t have peptidoglycan nor an outer membrane.

Do: They contain polysaccharide, protein, glycoprotein, or other complex macromolecules.

Tell me about the most common archaeal cell wall type. What is it? Why is it called as such? What is it made of? What are the three things it does?

S-layer (paracrystalline surface layer). It is called as such due to its somewhat regularly ordered structure. It is made of a layer of interlocking proteins or glycoproteins, (a) protecting against osmotic pressure, (b) keeping periplasmic proteins from floating way, and (c) blocking entry of large molecules, while being porous to small molecules.

What is pseudomurein? What is it made of? Where can you usually find this “pseudomurein”?

Archaeal cell wall type made of a complex peptidoglycan-like structure. Usually found in methanogenic Archaea (i.e. archaeal organisms making methane).

What are the six characteristics of pseudomurein that differentiates itself from a bacterial cell wall (CW)?

• N-acetyltalosaminuronic acid (NAT) instead of NAM.

• ß-1,3 bonds instead of ß-1,4 bonds.

• L-amino acids instead of D-amino acids.

• Nondegradable by lysozyme.

• Cross-link formation not inhibited by penicillin.

• If they have enough pseudomurein, may cause them to stain gram positive.

What are two groups of prokaryotes that do not have cell walls?

Mycoplasma (Bacteria) and Thermoplasma (Archaea).

What helps Mycoplasma survive without a cell wall?

They have sterols in their CM, adding structural integrity.

Note: Sterols are a group of lipids that give more support to the CM rigidity in the context of Mycoplasma.

Where would you find Thermoplasma? Why so?

Found in hot and acidic environments, they are extremophiles with a CM made of a tetraether lipid monolayer with its lipids linked to long heteropolysaccharides.

Note: A heteropolysaccharide is a polysaccharide containing different types of monosaccharides.