Lecture 3 Flashcards
What do reaction-diffusion mechanisms result in
Spontaneous pattern formation
What do spatiotemporal patterns emerge from
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.
Min oscillations - reaction diffusion system
+ loop - cooperative binding of MinD molecules.
- loop - MinE and MinD/C.
Diffusion of the activator plus delayed local activation of the inhibitor
Local inhibition of the activator
Vesicle transport
From Golgi to cell exterior; we also see transport from other organelles.
Microtubules
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.
Tubulin
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.
Tubulin polymerisation
Occurs more readily at the + end of the filament.
MT-associated proteins that regulate MT dynamics and organisation
gamma-TuRC, +TIPs, MAPs, MAP2, tau, kinesis-13.
Tau
Regulates binding of Mts and their stability, as well as the extent to which kinesis and dynein can bind MTs.
+TIPs
Remain associated with growing plus ends and can link them to other structures, such as membranes; preferentially associated with + ends of polymerising filaments
gamma-tubulin ring complexes
nucleate microtubule
polymerization at the centrosome; organise into interfacing web.
Microtubule motor proteins
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.
Speed of motor proteins
Usually median speed of 1 um/s
Does myosin transport material in axons
Experimental approach: Inject myosin-coated
bead into an axon. Watch under the microscope
to see if the beads move.
1) get some squid
In vivo reconstitution of motility
- 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
Fractionation methods
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.
