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Microtubule functions
- Cell shape
- Cell division
- Intracellular transport
- Cell motility
- Positioning of organelles
- Other specialized functions
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Marginal band
Microtubule structure in platelets that give the platelets their shape
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Best source of microtubules and why
Brain (neurons are highly asymmetric and have extensive microtubule network)
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How to purify microtubules
- Take brain tissue, make cold to decrease hydrophobic interactions between microtubules
- Spin in centrifuge, discard the pellet (organelles and other stuff)
- Add glycerol (water sequestering agent) and GTP to supernatant
- Increase temperature to allow hydrophobic interactions to occur again
- Spin, take the pellet (which is now the microtubules)
- Resuspend the pellet in cold temperature and repeat the cycle
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Structure of protofilament
- 13 subunits arranged around hollow core
- a- and b-tubulin heterodimer arranged in head to tail position
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Polarity of protofilament
- a- and b-tubulin heterodimers arranged in head to tail position
- + end at alpha end; - end at beta end
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2 examples of molecular motors from class and how they work
- Kinesins: anterograde transport (towards the + end)
- Dyneins: retrograde transport (towards the - end)
- Both work by hydrolysis of ATP for energy and "walk" along the microtubule
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Drugs that influence microtubule polymerization
- Depolymerizing (e.g. colchicine, nocodazole): depolymerize microtubules, inhibiting mitosis
- Polymerizing (e.g. vinca alkaloids, taxol): stabilize microtubule filaments and prevent normal spindle formation during mitosis
- For chemotherapy, give polymerizing drugs because the depolymerizing ones are too poisonous
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- end of microtubules is anchored on the ____
centrosome
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Centrosome
Main site of microtubule synthesis in the cell; where the - end is anchored
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Centriole structure
9 triplets of tubulin
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Exchangeability of GTP in microtubules
- Binding site of GTP on a-tubulin is non-exchangeable because the binding site is stuck between a- and b-tubulin (always GTP)
- Binding site of GTP on b-tubulin is exchangeable
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Theory of dynamic instability
- The reason why there is no simple equilibrium between tubulin heterodimers and microtubule polymers
- When GTP-bound tubulin heterodimer is at + end of growing polymer, GTP hydrolysis is triggered
- Tubulin is an enzyme
- If the rate of GTP hydrolysis exceeds the rate of addition of subunits, the cap will shrink and be lost, triggering depolymerization
- If the rate of GTP hydrolysis is slower than the rate of addition of subunits, the GTP cap will grow and the polymer will continue to grow
- In vivo, since the microtubule polymer is anchored to the centrosome at the – end, depolymerization can only go as far as the centrosome. And, in fact, since there are several GTP “caps” throughout a polymer (GTP hydrolysis is imperfect), it will only depolymerize back to the previous GTP cap
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