L18: Microbial Structures and Functions / Microbial Metabolism

Question 1

What is binary fission?

Answer 1

• Process that bacteria go through to produce an exact copy of genome (bacteria are haploid DS DNA molecule) and single cell divides into 2 daughter cells

Question 2

Describe growth curve phases of bacteria

Answer 2

1. ) Lag phase: period when bacteria are actively metabolizing and building up metabolites for cell division, no division has occurred yet
2. ) Exponential phase: period of division
3. ) Stationary phase: bacterial have used up all nutrients in media or have built up toxic metabolites – population size isn’t increasing
4. ) Decline phase: population is becoming reduced

Question 3

In terms of the growth curve phases, when are bacteria more or less sensitive to antimicrobial killing?

Answer 3

• Most sensitive during lag and exponential phase as active metabolic pathways can be targeted by the antimicrobials
• Less sensitive during stationary phase as bacteria have moved into dormant from vegetative state and are sporulating (in G+ bacterium only) – no active metabolism

Question 4

Minimum requirement for growth by bacteria

Answer 4

• C/N source, energy source, water and various ions

Question 5

Define aerobes, anaerobes and facultative anaerobes

Answer 5

• Aerobe: exclusively uses respiration to meet its energy need – therefore require oxygen to live
• Anaerobe: exclusively uses fermentation to meet its energy need – therefore don’t require oxygen to live
• Facultative anaerobes: can respire or ferment

Question 6

What are microaerophillic bacteria?

Answer 6

• Bacteria that grow best at low o2, but can grow without o2

Question 7

Why do obligate anaerobic bacteria die in oxygen?

Answer 7

• Oxygen is highly reactive when in forms such as hydrogen peroxide (h2o2) and superoxide anion (o2-). These organisms don’t have catalase and SOD to detoxify each of these respectively

Question 8

Enzymes that aerobes contain to deal with ROS?

Answer 8

• Catalase: breaks down H2o2 to h2o and o2
• SOD (superoxide dismutase): detoxifies o2- to h2o2 and o2
• Also peroxidase in obligate aerobes

Question 9

Two types of respiration

Answer 9

1. ) Aerobic: electron acceptor = molecular oxgen

2. ) Anaerobic: similar metabolism to aerobic respiration except nitrate or sulfate serve as terminal electron acceptors

Question 10

What is fermentation? Clinical relevance

Answer 10

• Anaerobic process by which organic metabolic intermediate derived from fermentable substrate serves as final electron acceptor
• Identification of bacteria can be done via end products of fermentation, unique to certain bacteria

Question 11

Describe efficiency of respiration vs fermentation

Answer 11

• Aerobic respiration is most efficient producing 19 x ATP than fermentation which is least efficient. Anaerobic respiration is somewhere in between

Question 12

Overarching targets of antibiotics

Answer 12

1. ) Cell wall synthesis (peptidoglycan)
2. ) DNA/RNA synthesis
3. ) Protein synthesis (against 50 or 30S ribosomes)
4. ) Antimetabolites

Question 13

How can antibiotics target nucleic acid synthesis in bacteria given humans also perform this synthetic pathway?

Answer 13

1. ) Targeting folate synthesis
   - Folate is essential in synthesis of purines (DNA and RNA) and thymidine (DNA)
   - Humans get folate from dietary sources, bacteria are required to synthesize it from para-aminobenzoic acid
   - Sulfonamides and DHFR inhibitors target enzymes in this synthetic pathway

2.) Bacterial DNA gyrase is required to unwind/win DNA: targeted by quinolones

Question 14

How can antibiotics target bacterial translation?

Answer 14

• Antibiotics against the 70S ribosomes (50 or 30S subunits). Humans have 80S ribosomes

Question 15

Describe peptidoglycan synthesis

Answer 15

1. ) NAG and NAM are activated by binding them to UDP
2. ) A pentapeptide is added to UDP-NAM
3. ) UDP-NAM-pentapeptide is attached to bacoprenol (lipid carrier on cytoplasmic portion of membrane) through pyrophosphate link, which releases UMP
4. ) NAG is added to NAM-pentapeptide bactoprenol complex
5. ) Bactoprenol carrier transports the complex across the membrane to outer membrane leaflet
6. ) Disaccharide unit is attached to growing peptidoglycan chain by enzymes called transglycosylases
7. ) Pyrophosphobactroprenol (empty) is converted back to bactoprenol and recycled, moved back to inner membrane leaflet
8. ) Transpeptidation occurs between free amine of diamino amino acid in third position of first peptidoglycan layer and D-ala at 4th position of other peptide chain releasing one of the D-ala on the other peptidoglycan layer. Occurs via transpeptidases (aka penicillin binding proteins – PBPs)

Question 16

What is the pentapeptide in peptidoglycan?

Answer 16

• 1: variable, 2: variable, 3: diamino amino acid (lysine and diaminopilemic acid), 4: D-ala and 5: D-ala

Question 17

Target of vancomycin

Answer 17

• Targets release of complex from bactoprenol on the outer membrane leaflet

Question 18

Target of bacitracin

Answer 18

• Prevents reclying of bactoprenol from outer leaflet to inner leaflet to continue peptidoglycan synthesis

Question 19

Target of penicillins and other beta-lactam antibiotics

Answer 19

• Targets transpeptidases and linking of 3rd diamino amino acid on one peptidoglycan layer to 4th D-ala on other peptidoglycan layer

Question 20

Target of cycloserine

Answer 20

• Prevents L-ala conversion to D-ala. D-ala is component of pentapeptide in peptidoglycan

Question 21

True / False. Peptidoglycan is a structure that is only found in bacteria.

Answer 21

• True

Question 22

During peptidoglycan formation, amino acids are added without the help of transfer RNA and ribosomes. True / False

Answer 22

• True

Question 23

In E.coli, crosslinking of peptidoglycan layers occurs between the amino acids
A.	D-ala and D-ala
B.	D-ala and L-ala
C.	D-ala and meso-diaminopimelic acid
D.	L-ala and meso-diaminopimelic acid
E.	L-ala and D-glutamic acid

Answer 23

• C.