Module 2 - Bacterial structure and function

Question 1

Binary fission: the process

Answer 1

Cell doubles in length, all other organelles are partitioned with an equal split on each side, septum forms, and then the cells fully split

Question 2

What stimulates cell division?

Answer 2

The FtsZ ring

Question 3

MreB: where is it present, what is it, and what shapes are the mutants?

Answer 3

Only present in rod-shaped bacteria

An actin homologue which forms a filamentous structure which acts as a scaffold for peptidoglycan synthesis machinery

Coccoid

Question 4

How does the insertion of new peptidoglycan work in rod-shaped bacteria?

Answer 4

Autolysin breaks the b1-4 linkage between NAM and NAG

Transglycosylase adds a new peptidoglycan unit

Question 5

How is more peptidoglycan formed in coccoid-shaped bacteria?

Answer 5

Since there is no MreB to synthesize the breakdown and synthesis of peptidoglycan, synthesis can only happen at the septum, when the cell divides

Question 6

The 4 phases of bacterial growth in culture

Answer 6

Lag phase, Log phase, Stationary phase, and Death phase

Question 7

Lag and Log phases: what are they and why do they occur?

Answer 7

Lag phase - The phase where there is no increase in the log number of viable organisms due to the bacteria adjusting to the new environment and growing

Log phase - The phase where there is an exponential growth of organisms. This usually forms a straight line.

Question 8

Stationary and Death phases: what are they and why do they occur?

Answer 8

Stationary phase - The phase where the culture size is stable and there are no more divisions due to lack of nutrients

Death phase - The phase where the culture begins to die out due to intolerable environment (build-up of toxic material, temp etc)

Question 9

Psychrophiles: what temps are their optimum?

Answer 9

<15°C

Question 10

Mesophiles: what temps are their optimum?

Answer 10

15-45°C

Question 11

Thermophiles: what temps are their optimum?

Answer 11

45-80°C

Question 12

Hyperthermophiles: what temps are their optimum?

Answer 12

> 80°C

Question 13

Examples of extremophile adaptations (psychrophiles)

Answer 13

1 - Unsaturated fatty acid lipid bilayer. This prevents the membrane from becoming waxy at low temps

2 - Altered proteins. More α-helices give enhanced flexibility

3 - Antifreeze molecules which bind ice crystals have been found in fish and other eukaryotes, but not in prokaryotes

Question 14

Examples of extremophile adaptations (thermophiles)

Answer 14

1 - Saturated fatty acid lipid bilayer. This remains stable at high temps

2 - Altered proteins. Fewer α-helices give more folding conformations to withstand heat

Question 15

Examples of extremophile adaptations (hyperthermophiles)

Answer 15

1 - No lipid bilayer. Phytane (C₄₀) hydrocarbon is present which creates a lipid monolayer

Question 16

How do bacteria cultures die out?

Answer 16

Not instantly - they die at a constant rate over time

Question 17

What steam sterilisation kills all known pathogens?

Answer 17

Temp - 121°C
Time - 20 mins
Pressure - 138 kPa

Question 18

Variety of bacterial cell shapes

Answer 18

Range from circular (coccus) to anything (ie can be stellar (star) shaped)

Question 19

What colours are Gram-positive and Gram-negative cells?

Answer 19

Positive - Purple
Negative - Pink

Question 20

Difference between the cell walls of Gram-positive and Gram-negative cells

Answer 20

Gram-positive cells have a thick layer of peptidoglycan as an outer membrane

Gram-negative cells have a thin peptidoglycan layer between the inner and outer membrane

Question 21

Gram stain process

Answer 21

Heat fix to raise cells to the top, crystal violet to dye the cells, fix with iodine to form a larger stained crystal, decolourise with ethanol/acetone, and counterstain

Both will be dyed purple after the crystal violet is used and the Gram-positive cells will remain purple throughout

Gram-negative cells will be dyed purple but then will be decolourised with acetone/ethanol and coloured pink with the counterstain

Question 22

The use of peptidoglycan in the cell wall

Answer 22

• The rigid structure of the wall gives the cell shape
• Protects from osmotic lysis
• Withstand high internal osmotic pressure
  (Gram-positive - 20 atm, Gram-negative - 0.3-3 atm)

Question 23

Strengths and weaknesses of peptidoglycan

Answer 23

Strength: extremely strong - boiled cells can have their intracellular components destroyed with the peptidoglycan bag remaining intact
Weakness: Antibiotics, lysozyme, and bacteriophage lysins

Question 24

Why can lacking peptidoglycan be beneficial?

Answer 24

Antibiotics won’t bind, protection value

Question 25

Peptidoglycan: what is its basic structure?

Answer 25

Altering amino acid chains (N-acetyl glucosamine (NAG) and N-acetyl muramic acid (NAM)) with glycosidic bonds binding them at the top and bottom of the peptidoglycan. The vertical gap between the top and bottom is filled with amino acids

Glycan is conserved highly, peptides may be variable

The backbone carbohydrate sugar is around 10-65 sugars per chain

Question 26

How is peptidoglycan bonded in the cell wall of bacteria?

Answer 26

Vertical bonding - peptide bonds which bind the amino acids

Horizontal bonding - glycosidic bonds

Question 27

Gram-positive cell wall structure

Answer 27

Homogenous architecture vertically filled with large amounts of glycine with peptidoglycan (PG) and teichoic acid (TC, a polymer of glycerol (3C) and ribitol (5C) phosphate)

Question 28

Gram-negative cell wall structure

Answer 28

Heterogenous architecture with an outer membrane, periplasm on both sides of the peptidoglycan, and an inner membrane

Contains lipopolysaccharides

Question 29

Lipopolysaccharides: what are they and what do they consist of?

Answer 29

Complex glycolipids that are exclusively present in the outer leaflet of the outer membrane of Gram-negative bacteria which renders outer membrane asymmetric and less fluid

LPS consist of:
* Conserved hydrophobic membrane anchor - lipid A
* Variable oligosaccharide core region
* Highly variable O-antigen polysaccharide forming a repeating oligosaccharide

Question 30

Endotoxin: what is it and what can it cause to occur?

Answer 30

Stimulates the innate immune system to trigger an inflammatory response which can cause the body to go into endotoxin shock which can lead to multi-organ failure and death

Endotoxin = extracellular Lipid A

Question 31

Archaeal cell envelope - extremophile

Answer 31

Contains an S-layer of two proteins and membrane anchoring proteins

Tetra-ethyl lipids in the lipid monolayer (also impermeable to protons)

Question 32

Archaeal cell envelope - mesophile

Answer 32

S-protein layer above pseudomurein which is above a membrane of diether lipids

Question 33

Archael pseudomurein

Answer 33

Essentially a substitute for peptidoglycan

Some may have it (methanogens) but some may not

Altering amino acid chains (N-acetyl glucosamine (NAG) and N-acetyl talosaminuronic acid (NAT)) with glycosidic β1-3 bonds (which are resistant to lysosome degradation) binding them at the top and bottom of the pseudomurein. The vertical gap between the top and bottom is filled with amino acids (only L-amino acids present, no D-forms)

Question 34

Surface layers

Answer 34

The outer membrane of archeal cells which is mostly composed of a paracrystalline protein layer

Question 35

What types of arrangements can flagella have?

Answer 35

Monotrichous - one
Lophotrichous - many on one side
Amphitrichous - one each side
Peretrichous - many on every side

Question 36

Flagella mediated motility

Answer 36

Flagella are semi-rigid and rotate up to 300 revolutions per second

Bacterial cells swim at 0.00017 km/h but this is 50-60 cell lengths per second

For comparison, Cheetah runs at 110 km/h or 25 body lengths per second

Question 37

Flagella ultrastructure

Answer 37

Filament - acts as the propellor
Hook - joins filament and basal membrane

Hook’s structure:

L ring, P ring, MotB, MS, MotA ring, and C ring

Question 38

The flagella’s hook’s structure

Answer 38

• L ring - in line with the outer membrane
• P ring - in line with peptidoglycan
• MotB - in the periplasm below the peptidoglycan
• MS - in line with the inner membrane, top of the machinery that moves the flagella
• MotA - in line with the inner membrane, a channel for protons
• C ring - Inside the inner membrane, the bottom of the machinery that moves the flagella

Question 39

What facilitates flagella movement

Answer 39

The rotation of the C and MS rings because of MotA, causing the hook and filament to move.

The P and L rings do NOT move at any point (and are not present in Gram-positive bacteria)

Question 40

Flagella: what are the types of flagella movement and how does the motility work?

Answer 40

Phototaxis and chemotaxis

CCW (counterclockwise - flagella bundle to move in one direction) to move in one direction and CW (spread out flagella) to stop moving

Question 41

Fimbriae and pili on the bacterial cell surface: what are they, what are their functions, and what on earth is type IV pili and what does it do?

Answer 41

Thin (2-10 nm wide) proteinaceous and filamentous structures that project from the cell surface

Fimbriae are often peritrichous and enable commensals and pathogens to adhere to surfaces and tissues (often via adhesins at the tip to counteract sheer forces), obtain nutrients at specific niches, and form pellicles/biofilms at interfaces/surfaces

Pili have a similar structure (though fewer and more extended than fimbriae) and they allow adherence to host tissues (ie E. coli mannose receptor on pili tip), exchange of genetic material (conjugation pili)

Type IV pili – transformation, twitching motility on a surface, and adherence essential in pathogens such as G-ve’s Vibrio cholerae, Neisseria gonorrhoeae and Streptococcus pyogenes (G+ve), also found in Archaea

Question 42

E.Coli: the different types of P-type and what do each of them do?

Answer 42

• PapG - adhesin
• PapF - adaptor protein
• PapE - fibrillar protein
• PapK - adaptor protein
• Pap A - structural subunit approx.1000 copies
• PapC - assembly/secretion of subunits