Lecture 3

Question 1

What do reaction-diffusion mechanisms result in

Answer 1

Spontaneous pattern formation

Question 2

What do spatiotemporal patterns emerge from

Answer 2

Amplification of stochastic fluctuations - in the concentration of either of these diffusible species.

Activatorandinhibitor concentrations
fluctuate randomly. Positive feedback loop amplifies random increases in activator concentration

[ ] of the activator will be slightly higher; can get travelling waves, global oscillation patterns.

Question 3

Min oscillations - reaction diffusion system

Answer 3

+ loop - cooperative binding of MinD molecules.

• loop - MinE and MinD/C.

Question 4

Diffusion of the activator plus delayed local activation of the inhibitor

Answer 4

Question 5

Local inhibition of the activator

Answer 5

Question 6

Vesicle transport

Answer 6

From Golgi to cell exterior; we also see transport from other organelles.

Question 7

Microtubules

Answer 7

Polar; formed from the polymerisation of tubulin dimers (beta- and alpha-; beta oriented towards plus end, alpha towards negative).
Have an outer diameter of 25nm. Long, straight, frequently have one end attaached to MTOC - centrosome.

Built from 13 protofilaments - linear strings of tubulin joined end to end, that associate to form a hollow cylinder.

Easier to lose a subunit at beginning of end of the filament than the middle because you’d have to break more bonds = can resist thermal breakage while having rapid subunit additional and removal at the filament ends.

Question 8

Tubulin

Answer 8

Compact, globular. Form as helical assemblies of subunits = microtubules. Self associate using combination of end-to-end and side-to-side protein contacts. Held together by weak non-covalent bonds.
Asymmetrical, bind to one another head-to-tail so all point in one direction.
Tubulin is an enzyme that catalyses the hydrolysis of GTP which enables MTs to remodel rapidlyy.

Question 9

Tubulin polymerisation

Answer 9

Occurs more readily at the + end of the filament.

Question 10

MT-associated proteins that regulate MT dynamics and organisation

Answer 10

gamma-TuRC, +TIPs, MAPs, MAP2, tau, kinesis-13.

Question 11

Tau

Answer 11

Regulates binding of Mts and their stability, as well as the extent to which kinesis and dynein can bind MTs.

Question 12

+TIPs

Answer 12

Remain associated with growing plus ends and can link them to other structures, such as membranes; preferentially associated with + ends of polymerising filaments

Question 13

gamma-tubulin ring complexes

Answer 13

nucleate microtubule
polymerization at the centrosome; organise into interfacing web.

Question 14

Microtubule motor proteins

Answer 14

kinesin-1 (+ end directed) and dynein (- end directed).
They have a range in speed and ATP hydrolysis rates.

The motor proteins kinesin and dynein hydrolyze ATP in order to
transport cargo along microtubules.

Question 15

Speed of motor proteins

Answer 15

Usually median speed of 1 um/s

Question 16

Does myosin transport material in axons

Answer 16

Experimental approach: Inject myosin-coated
bead into an axon. Watch under the microscope
to see if the beads move.

1) get some squid

Question 17

In vivo reconstitution of motility

Answer 17

1. centrifuge such that soluble proteins are separated from ATP, membrane organelles, and MTs.

Experiment one: combine MTs, ATP, membrane organelles, pipette onto microscopic slide. ==> NO movement.

Experiment 2: Combine the above with soluble proteins as well. Add to microscopic slide. MOVEMENT.

Experiment 3: the negative control. Mts, ATP, soluble proteins. MOVEMENT. Motors on the glass slide the MTs

Question 18

Fractionation methods

Answer 18

Centrifugation - separating molecules by densities.

Column chromatography - separating molecules according to polarity or hydrophobicity. Principle of Adsorption, in which a mixture of components dissolved in the mobile phase is introduced in to the column and the components move depending on their relative affinities. The choice of the solvent depends on the solubility characteristics of the mixture.