User:Andy Maloney/Notebook/Lab Notebook of Andy Maloney/2009/04/21/Kinesin & Microtubules

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Below are my interpretations on the article I've read. Please don't use my info as the correct interpretation without reading the paper first.


How calcium causes microtubule depolymerization


There are lots of good and interesting things in this paper. I will list most of them below.

  • Experiment
    • They used calcium to trigger catastrophes in microtubules. Catastrophes mean that microtubules shrink. Not sure why this terminology is used.
    • Two types of microtubules were investigated
  1. phophocellulose-purified (PC) microtubules
  2. Microtubules with microtubule associated protein (MAPs) on them.
  • They show that calcium induces the disassembly of microtubules for both PC and MAP microtubules.
  • They showed that rescue rates reduced when the temperature of the microtubule solution decreased. Rescue is when microtubules start to grow again.
Koch question: Will we need to get a temperature stabilizer for our experiments?
  • Question: This paper shows that free calcium will affect microtubules. What about calcium affecting microtubules fixed with Taxol?
  • They showed that above 500 µM of free calcium, catastrophe rates of microtubules were enough to make them shrink.
  • They also note that free magnesium will cause catastrophe as well. But, it starts at 5 mM.
  • Note: Lots of buffers I've seen people use include using MgCl2 and Casein. Casein has calcium phosphate in it and as I just found out, magnesium will cause catastrophe as well. This is probably why people use EGTA/EDTA in their buffers.
    • Question: Is there a specific reason as to why people use MgCl2 in their buffers? Does either kinesin or microtubules need a little bit of it to work? Yes! Apparently someone has asked this question and this paper talks about it. Magnesium actually aids in microtubule growth.
  • They note that MAP microtubules are more stable and exhibit better contrast in their DIC images.
  • Of course, the most interesting part of the paper doesn't have data showing this. Apparently they saw MAP microtubules stop all rescue/catastrophe events after about 20 - 30 minutes at 37˚C. The microtubules were even stable with the addition of mM concentrations of calcium. Whether or not this statement is true needs to be investigated.
  • They talk a lot about GTP stabilization and GTP caps on microtubules.
    • Note: Should I be concerned about knowing what this means?

Take home

Calcium affects microtubules. Casein has calcium in it. People use EGTA/EDTA to stop calcium from screwing with microtubules but, does calcium screw with microtubules fixed with Taxol?

Good paper.