9 - BACTERIAL MOTILITY AND TAXIS

Question 1

Tactic response requires:

Answer 1

1) Motility - swimming

2) Sensory mechanism

Question 2

Various forms of taxis:

Answer 2

Chemotaxis, aerotaxis, phototaxis, thermotaxis, pH taxis, magneto taxis

Question 3

Different flagella arrangements:

Answer 3

Polar flagella (Vibrio Cholera, pseudomonas), peritrichous flagella (E.coli)

Question 4

Basic components of flagella and their sizes:

Answer 4

Basal body, hook and filament - 10-15 um long, 14nm diameter

Question 5

What is the filament made out of?

Answer 5

FliC, flagellen

Question 6

What is the hook made of?

Answer 6

FlgE and hook associated proteins (HAPs) e.g. HAP2 - cap protein on growing filament - will move along as filament is made

Question 7

How many different proteins make up the basal body?

Answer 7

26 different proteins

Question 8

Where is FliC made?

Answer 8

Made in cell, and polymerises and assembles on the cell surface

Question 9

What different rings are part of the basal body and where are they located ?

Answer 9

L-ring, P-ring, MS-ring, and C ring.
L-ring = OM absent in g+ bacteria
P-ring = contacts PG layer
MS-ring = in CM 
C-ring = in cytoplasm

Question 10

Which protein confer motor function?

Answer 10

Motor proteins (MotA and MotB) and switch proteins

Question 11

How many mot proteins are there, and where are they located?

Answer 11

11 mot proteins, surrounding the MS ring

Question 12

What does in motor protein unit consist of?

Answer 12

MotA4 and MotB2 = 1 unit

Question 13

What protein does Mot interact with?

Answer 13

FliG

Question 14

What proteins form the C ring? How many copies of the protein?

Answer 14

Switch proteins - FliG, FliC, FliN, FliM - 26 copies

Question 15

Components that rotate and components that do not:

Answer 15

Rotate: C ring, MS ring, Rod-Hook-Filament

Do not Rotate: Mot proteins (P ring L ring)

Question 16

Which aa in MotB are critical for rotation?

Answer 16

aa in MotB - Asp32

Question 17

What chemical property plays a role rotation on aspartate residues in motB?

Answer 17

Dicarboxylic acid - 2 COOH groups, one in peptide bond and one can be protonated and has a role in pmf and generating rotational force

Question 18

Other that aspartate what amino acids are involved in rotation in motB?

Answer 18

Proline173 and Proline22, mutation causes reduced rotation. Proline involved in rotation as can undergo conformational change

Question 19

When aspartate is protonated it leads to a conformational change which is then transmitted to which protein?

Answer 19

Switch protein FliG, which then transmitted to the next switch protein in the MS ring - this is what generates rotational movement and force

Question 20

When peritrichous flagella rotate counter clockwise how do bacteria move?

Answer 20

Smooth swimming

Question 21

When peritrichous flagella rotate clockwise how do bacteria move?

Answer 21

Tumbling mode - not swimming smoothy - jittery motion on the spot

Question 22

Smooth swimming and tumbling cause the bacteria to move in a…

Answer 22

Random walk - switch between tumbling and smooth swimming

Question 23

What causes bias random walk?

Answer 23

Chemical in the environment e.g. glucose

Question 24

What changes during a bias random walk?

Answer 24

Extended time smooth swimming, more CCW rotation of flagella. Shows purposeful behaviour.

Question 25

What is chemotaxis?

Answer 25

Purposeful behaviour to or from a compound

Question 26

Describe experiment to show bias random walk/chemotaxis:

Answer 26

Capillary tube containing: 1) nutrient rich media 2) same nutrient poor medium as in growth vessel 3) toxin was placed in a culture vessel containing nutrient poor medium. Then a viable cell count was taken.

Question 27

What were the results of the capillary tube experiment o show evidence of chemotaxis?

Answer 27

Significantly more bacteria above control (same nutrient poor media) when nutrient rich media in capillary tube, significantly less bacteria when toxin in tube. This shows bacteria must be displaying purposeful chemotaxis behaviour AND SENSING concentration.

Question 28

What does the frequency of tumbling depend on?

Answer 28

The frequency of tumbling depends on change in concentration of the stimulus

Question 29

What are two possible sensing mechanism?

Answer 29

1) Spatial sensing - sense change between front and back - discounted too small distance, change in conc would be very small 2) Temporal sensing - change in concentration of attractant or repellent with time cause switch between tumbling and smooth swimming behaviour

Question 30

Mechanism of temporal sensing requires:

Answer 30

1) memory - compare current and past measures 2) adaptation mechanism - adapt to high concs in order to detect higher ones - adaption is fundamental * respond to change not absolute conc

Question 31

What are MCPs?

Answer 31

Methyl accepting chemotaxis proteins. They span the CM. Different bacteria have different MCPs as different environments require different detection.

Question 32

How many MCPs does E.coli have?

Answer 32

4 Tar, Tsr, Trg, Tap

Question 33

What does Tar sense?

Answer 33

Attractant: Asp Maltose Repellent: Co Ni

Question 34

What does Tar sense?

Answer 34

Attractant: Serine Repellent: Acetate, benzoate

Question 35

What does Trg sense?

Answer 35

Attractant: Galactose, ribose

Question 36

What does Tap sense?

Answer 36

Dipeptides

Question 37

What domains do MCPs have?

Answer 37

Sensing domain - periplasm | Signalling domain - cytoplasm

Question 38

Ternary Complex:

Answer 38

MCP, CheW, CheA

Question 39

Stabilised Ternary Complex occurs under what condition?

Answer 39

Forms under no attractant.

Question 40

Stabilied Ternary Complex

Answer 40

No attractant causes CheA activity - autokinase activity phosphorlates histidine by hydrolysing ATP. Then transfers P from CheA to CheY (A key signalling molecule). CheY-P binds to switch protein in C ring = flagella rotation CW and tumbling behaviour. - So if no attractant = increased chance of tumbling (not binary)

Question 41

Ternary complex becomes destabilised under what conditions

Answer 41

Attractant bound = destabilised

Question 42

How is ternary complex destabilised

Answer 42

No longer mechanism to make CheY-P . CheZ dephosphorylates an existing CheY CheY-P no longer interacts with switch proteins. No CW rotation of flagella Increased CCW rotation and increase swimming

Question 43

How does bacteria know to keep moving toward something?

Answer 43

1st time interact = ternary destabilise and move toward How does bacteria know to keep moving toward something? - memory - adaption

Question 44

Adaption mechanism

Answer 44

CheR is a methyl transferase and transfers Me to exposed glutamic residues on MCP. Glutamic residue exposed due to attractant binding. Methylation = restabilisation of ternary complex formation - switch back to no attractant - bacterium now adapted to concentration To destabilise ternary complex needs to encounter increased conc of attractant.

Question 45

How to reset adaption mechanism of chemotaxis if go from high to low conc?

Answer 45

RESET IF BACTERIA MOVE TO LOWER CONC | methyl esterase CheB, removed Me from MCP, this destabilises ternary complex = attractant bound state = swimming.

Question 46

Are the MCP evenly distributed?

Answer 46

Not entirely, seem to be more at 'front' end opposed to flagella end. Spatial separation between detection and activation