User:Andy Maloney/Kinesin & Microtubule Page/Kinesin papers/Effect of temperature on kinesin driven microtubule gliding and kinesin ATPase activity

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Disclaimer

These are my notes on the below paper. Please do not read my notes until you have read the paper.

Paper

Effect of temperature on kinesin-driven microtubule gliding and kinesin ATPase activity

Notes

  • This paper talks about the changes in gliding motility speeds due to temperature effects.
  • They used porcine brain kinesin that they purified themselves.
  • They purified tubulin from porcine brains as well. They polymerized the microtubules in 10 µM Taxol.
  • Their buffer consisted of:
    • 50 mM imidazole
    • 0.5 mM MgCl2
    • 0.5 mM EGTA
    • 0.1 mM DTT
    • pH 6.8
    • They added 100 mM NaCl, 0.5 mM disodium ATP, and 10 µM Taxol for motility.
  • Their final concentration of tubulin and kinesin were:
    • Kinesin 70 µg/mL
    • Tubulin 40 µg/mL
  • They incubated microtubules and kinesin for 10 minutes before adding them to a slide.
  • They used 5 mg/mL BSA as their passivator.
  • They used the Argus 20 image processing software.
  • The take home from this paper is that speed from gliding motility assays are dependent on the temperature.
Steve Koch 23:34, 3 March 2010 (EST): One interesting thing I noticed upon re-reading. At Sandia we discovered that not only did thermomyces kinesin aggregate at 42C, but so did drosophila kinesin. Given the unique way in which they prepared their assays, I think they incubated the kinesin at high temperature in solution before putting on surface (Figure 3) ... so aggregation of kinesin likely accounts for the inactivity. As for figure 1, I don't know if the high temp behavior (plateau) is explained by the same thing.