3 - The Cell Wall

Question 1

Functions of cell wall

Answer 1

• Provides shape
• Helps anchor flagella
• Prevents cell bursting
• May contribute to pathogenicity

Question 2

Cell wall structure

Answer 2

• Composed of peptidoglycan (murein)
• Mesh like polymer enclosing the cell wall (peptidoglycan sacculus)
• Repeating NAG and NAM molecules in a chain

Question 3

What does NAG stand for

Answer 3

N-acetylglucosamine

Question 4

What does NAM stand for

Answer 4

N-acetylmuramic acid

Question 5

What bond are sugars linked by in cell wall

Answer 5

• Beta 1, 4, glycosidic bonds
• Sugar chains are cross linked by tetrapeptides (4 alternating D- and L- amino acids, connected to NAM)

Question 6

Peptidoglycan cross links

Answer 6

• The presence of D-amino acids protects against most peptidases which recognise only L-amino acids in proteins
• Three of the amino acids are not found in proteins: D-glu, D-ala, mesodiaminopimelic acid (DAP)

Question 7

Peptidoglycan sacculus

Answer 7

• Strong but elastic
• Able to stretch and contract in response to osmotic pressure

Question 8

Gram positive cell wall

Answer 8

• Comprises many peptidoglycan layers forming a dense network
• Cell wall is negatively charged
• In gram stain, ethanol shrinks the pores of thick peptidoglycan, thus crystal violet is retained and stains bacteria purple

Question 9

Why is cell wall negatively charged

Answer 9

due to peptidoglycan amino
acids and phosphate groups on teichoic acids

Question 10

Lipoteichoic acids

Answer 10

Covalently bound to plasma membrane lipids (anchor wall to PM)

Question 11

Functions of teichoic acids

Answer 11

• Taxonomically useful
• Maintain cell envelope structure by anchoring wall to PM
• Some involved in binding pathogenic bacteria to host tissues
• Help protect cell from harmful chemicals (e.g. antibiotics)
• Function in ion uptake and cell division processes

Question 12

Gram negative cell wall

Answer 12

• Does not contain teichoic acids
• Located in periplasmic space (periplasm)
• Has one or a few layers of peptidoglycan
• Not as highly cross linked as gram +’ve PG (larger pores, crystal violet removed and counterstain pink)

Question 13

Where does cell wall synthesis occur

Answer 13

The cytoplasm, plasma membrane and periplasm

Question 14

Steps of cell wall synthesis in the cytoplasm

Answer 14

1. UDP derivatives of NAM and NAG are made
2. Sequential addition of amino acids to UDP-NAM to form NAM-pentapeptide (has additional D-alanine)

Question 15

Steps of cell wall synthesis in plasma membrane

Answer 15

3. NAM-pentapeptide is transferred to BP phosphate via pyrophosphate bond
4. UDP transfers NAG to BP-P-P-NAM-Pentapeptide
5. Bactoprenol carrier transports completed NAG-NAM-pentapeptide unit across the membrane

Question 16

How does NAG-NAM pentapeptide cross the membrane

Answer 16

• Flipped across by flippase enzyme
• BP-P-P remains embedded in the membrane

Question 17

Steps of cell wall synthesis in the periplasm

Answer 17

6. NAG-NAM-pentapeptide is attached to the peptidoglycan
   chain, increasing its length by one repeat unit (transglycosylation)
7. Bactoprenol carrier moves back across membrane, losing a phosphate, ready to begin a new cycle (BP-P-P –> BP-P)
8. Peptide cross-links between glycan chains are formed by transpeptidation, mediated by transpeptidases

Question 18

Transpeptidation

Answer 18

Terminal alanine removed to
give a tetrapeptide (TP): this
provides energy to link the
peptides

Question 19

Antibiotics which inhibit cell wall synthesis

Answer 19

Penicillins (e.g. penicillin, ampicillin, methicillin) and cephalosporins (e.g. cephalothin)

Question 20

how do antibiotics which inhibit cell wall synthesis work

Answer 20

• Block murein cross-linking by binding to transpeptidases
• β-lactam ring resembles D-alanine-D-alanine in murein pentapeptide
• Transpeptidase binds to penicillin instead of its true substrate

Question 21

Why do β-lactams cause cell lysis

Answer 21

• Penicillins (PC) act only on growing bacteria
• Penicillin binding proteins (PBPs) first noted capacity to bind PC
• PBPs include transpeptidases, and some autolysins
• Penicillins bind to PBPs and may thus destroy bacteria by disregulating the autolysins

Question 22

What are autolysins

Answer 22

• Enzymes that hydrolyse bonds in existing PG to make room for new units e.g. endopeptidases
• Autolysins normally function in well-regulated way to allow cell growth and division
• Autolysin activity is tightly controlled as unregulated activity would weaken cell wall

Question 23

Where is lysozyme found

Answer 23

Sweat glands, mucous membranes (eyes, respiratory, digestive and urogenital tracts)

Question 24

How does lysozyme attack gram positive cell wall structure

Answer 24

• Breaks β-1,4 glycosidic bonds (NAM-NAG)
• Lysozyme-treated Gram positive cells form protoplasts

Question 25

Protoplasts

Answer 25

• Spherical
• Osmotically sensitive cells, bounded only by cell membrane.

Question 26

How does lysozyme attack gram negative cell wall structure

Answer 26

• Breaks β-1,4 glycosidic bonds (NAM-NAG)
• Form spheroplasts
• EDTA treatment plus l

Question 27

Spheroplasts

Answer 27

• Spherical osmotically-sensitive cells
• Bounded by PM but retaining fragments of OM and PG.
• Damage to OM is required to form them, e.g. by EDTA.

Question 28

L forms

Answer 28

• Protoplasts and spheroplasts which can grow/divide
• Penicillins can also cause formation of L forms
• Responsible for some
  chronic hard-to-treat infections

Question 29

Why are L forms responsible for hard to treat infections

Answer 29

• They are resistant to wall-acting antibiotics
• May persist in the body and revert to cell wall-production once treatment is ended